} Jeol sells a high resolution standard (gold on carbon) that we would like } to purchase. Does anyone have info: (part number, price, where to order)? } Many thanks. } } #################################################################### } John J. Bozzola, Ph.D., Director } Center for Electron Microscopy } Neckers Building, Room 146 - B Wing } Southern Illinois University } Carbondale, IL 62901-4402 } U.S.A. } Phone: 618-453-3730 } Fax: 618-453-2665 } Email: bozzola-at-siu.edu } Web: http://www.siu.edu/departments/shops/cem.html } ####################################################################
Most (all) of the EM supplies companies sell these types of specimens - Agar, SPI, Ted Pella, etc. I'd check their prices as well, since you might find them cheaper than EM manufacturers.
More generally, how do others find this specimen performs for resolution checks, particularly at very high resolution and/or low kV? Isn't the carbon a potential/real source of contaminations? I've seen tin spheres on aluminium being promoted as a resolution test specimen without the contamination problem. Is this a good substitute?
Regards,
--------------------------------------------------------------- Dr L. P. Stoter Technical Editor, MICROSCOPY & ANALYSIS Technesis 17, Rocks Park Road email: LPS-at-teknesis.demon.co.uk Uckfield, E. Sussex Phone: +44 (0)1825 766911 TN22 2AT Fax: +44 (0)1825 766911 United Kingdom ---------------------------------------------------------------
I have a turbopump horror story with a happy ending! A number of years ago, we decided to upgrade the diff pump which came on our Etec with a turbo. At that time greased bearing turbos were just appearing on the market. We ordered and installed a 360 l/s pump in place of the diff unit. The Etec e-optics system was quite susceptible to vibration which proved to be a problem, limiting effective maximum mag to between 5-10kX. Worse yet was the reliability. I have forgotten exactly how many (warranty and otherwise) pumps we went through. Must have been more than 5-6 in just a year or two. Bearing failure was the culprit. Some pumps would last for months, others failed only a few hours after installation. { {Greased bearing pumps have since improved, I hear} } Then the maglev pump hit the market! We replaced the ball bearing unit with a Leybold 340 l/s maglev turbo. That was somewhere over eight years ago. The system has been crashed to atm several times (valve failure) and been shut down quite a
number of times. It is still going strong with no problems. ...Love it. Must admit the Etec vacuum valving arrangement allows the pump to run continuously - it is not shut down for sample changes.... As far as the vibration problem, I can't tell I have a turbo pump on the system. The only drawback was the cost of the maglev pump. Given the maintenance free, vibration free, clean vacuum, it was worth it.
Good morning: Once again, I have a bit of a puzzle. I'd like to know _specifically_ the mechanism that generates continuum X-rays or "Bremstralung". There is variation among my text references as to the precise nature of the event that produces this signal. Several texts state that the interaction is inelastic and results from the slowing of electrons as they pass near the nucleus (electron-nucleus interaction), while others state that it is, again, inelastic, and is an primary electron-outer shell electron interaction (specific references are not given to avoid finger-pointing). My understanding of the term "inelastic" is that it refers to electron scattering of primary (beam), backscattered, and secondary electrons, and not to electron-atomic nucleus interaction. If someone could take the time to explain in gory detail, I'd appreciate it very much. I have a list of people to thank and a summary of my Holey grid responses to post, but that will come later. Many thanks for enlightenment. Interactively yours, Dwight
Dwight Beebe E-mail: beebed-at-ere.umontreal.ca Institut de recherche en biologie vegetale Voice: 514-872-4563 Universite de Montreal FAX: 514-872-9406 4101, rue Sherbrooke est Montreal, Quebec H1X 2B2 Canada
At 10:04 AM 4/1/97 -0400, Dwight Beebe wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
I don't believe we'll ever know the specific mechanism but the macroscopic model we have is apparently built upon an "apparent" bi-modal distribution of "events" we describe as "elastic" and "inelastic", and the (simple) definitions as I interpret them are: "If energy is transferred, then call it inelastic" ... "If very little energy is tranferred, then call it elastic". Another definition of "inelastic" would claim that characteristic information should be the result of an inelastic event.
(... an example of characteristic info gained would be a characteristic x-ray or a auger electron, whereas a backscattered electron, while informative, is not characteristic and is the result of an elastic event ...)
Since the x-ray contiuum is obviously the result of energy transfer it comes under the inelastic event catagory ... yet while it offers no characteristic information (like BSE, at least macroscopically) it has to be attributed to a "continuum" of energy transfers and while we can't "see" what actually goes on (orbital vs. nuclear), why claim one or the other???
... my $0.02 ...
cheerios, shAf
{\/} /\ {\/} /\ {\/} /\ {\/} cogito, ergo ZOooOM {\/} /\ {\/} /\ {\/} /\ {\/} Michael Shaffer, R.A. - University of Oregon Electron Probe Facility mshaf-at-oregon.uoregon.edu -or- mshaf-at-darkwing.uoregon.edu http://darkwing.uoregon.edu/~mshaf/
} I am interested in HREM image simulations of III-V semiconductors and I am } looking for any references where I can find the values of the Debye-Waller } temperature factors for such atoms as P, Ga, IN, As etc. } I would be extremely grateful for any helpful information
} Sincerely yours
} Rafal Spirydon
} Dept. of Materials Science and Engineering } Kwangju Institute of Science and Technology } South Korea
A good source of information is 'Debye-Waller Factors of Zinc-Blende-Structure Materials -- A Lattice Dynamical Comparison' by John S. Reid, Acta Cryst. (1983) A 39, 1-13. This contains calculated Debye-Waller Factors for the two atom species for various materials at a range of temperatures.
Another good place to find calculated ELEMENTAL Debye-Waller factors is 'Debye-Waller Factor for Elemental Crystals' by V. F. Sears and S. A. Shelley, Acta Cryst. (1991) A 47, 441-446. Alternatively, Sears and Shelley's results have been tabulated in 'Debye-waller Factors and Absorptive Scattering Factors of Elemental Crystals' by L.-M. Peng, G. Ren, S. L. Dudarev and M. J. Whelan, Acta Cryst. (1996) A 52, 456-470.
My own experience suggests that all of these sources are reasonably accurate (where, for Debye-Waller factors, 'reasonably' probably means less than ~10%-20% error). However, it still leaves you with the problem of deciding exactly what the temperature of your sample is under the electron beam (especially if you've cooled the sample to liquid nitrogen temperatures)!!
Regards,
Martin Saunders, Center for Materials Science and Engineering, Department of Mechanical Engineering, Naval Postgraduate School, Monterey, CA 93943, USA.
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Colleagues: Is anyone familiar with Sequenza staining racks? These are used to hold microscope slides for staining, and I'm not sure how they are different from your basic, run-of-the-mill slide racks. I haven't been able to find them in any catalogs.
TIA, Bev Maleeff
SmithKline Beecham Pharmaceuticals Toxicology-US, UE 0462 709 Swedeland Road King of Prussia, PA 19406 610/270-7987 610/270-7202 fax Beverly_E_Maleeff-at-sbphrd.com
} Good morning: } Once again, I have a bit of a puzzle. I'd like to know } _specifically_ the mechanism that generates continuum X-rays or } "Bremstralung". There is variation among my text references as to the } precise nature of the event that produces this signal. Several texts state } that the interaction is inelastic and results from the slowing of electrons } as they pass near the nucleus (electron-nucleus interaction), while others } state that it is, again, inelastic, and is an primary electron-outer shell } electron interaction (specific references are not given to avoid } finger-pointing). My understanding of the term "inelastic" is that it } refers to electron scattering of primary (beam), backscattered, and } secondary electrons, and not to electron-atomic nucleus interaction. If } someone could take the time to explain in gory detail, I'd appreciate it } very much. } I have a list of people to thank and a summary of my Holey grid } responses to post, but that will come later. Many thanks for } enlightenment. } Interactively yours, } Dwight } } } Dwight Beebe E-mail: } beebed-at-ere.umontreal.ca } Institut de recherche en biologie vegetale Voice: 514-872-4563 } Universite de Montreal FAX: 514-872-9406 } 4101, rue Sherbrooke est } Montreal, Quebec H1X 2B2 } Canada
This is going back a few years, but .....
The "Bremstralung" originates ONLY because the electrons are accelerated (or deaccelerated). The mechanism that causes the change in velocity is irrelevant. The cause comes from Special Relativity, as applied to charged particles - change the velocity of a charged particle and it will radiate.
The wavelength(?) and intensity(?) of the radiation will depend on factors like the change in velocity, mass and velocity of particle. The effect is only significant at near-relativistic velocities. Note also, it is specifically a change in VELOCITY, a vector quantity, not speed. You see a similar effect in sychrotrons, where the speed of the charge particle is constant but its direction is changed, so resulting in sychrotron radiation.
Additionally, don't mistake the background you see from an EDX/WDX detector on an electron microscope. Some of its characteristics are related to Bremstralung, but, especially at the low energy end, the overall response of the detector and amplifier are much more significant.
Regards, Larry Stoter
--------------------------------------------------------------- Dr L. P. Stoter Technical Editor, MICROSCOPY & ANALYSIS Technesis 17, Rocks Park Road email: LPS-at-teknesis.demon.co.uk Uckfield, E. Sussex Phone: +44 (0)1825 766911 TN22 2AT Fax: +44 (0)1825 766911 United Kingdom ---------------------------------------------------------------
MSA Recipients; I have a LEKTRA "Densi-timer" model PTM-4A which is in need of repair or replacement, but the company is no longer in business. The Densi-timer is a three-piece light meter for black and white only photo enlarging which reads an exposed area on the easel/paper to determine exposure time based on paper characteristics, rheostat setting, and magic. Any assistance in repair or suggestions for replacement will be greatly appreciated. Thanks. Merci. Shukran. Gracias. Danke. ------------------------------------- Name: Winston Wiggins Carolinas HealthCare System Charlotte, NC USA E-mail: wwiggins-at-carolinas.org Fax: 704-355-7648 Voice:704-355-7220
Message-Id: {199704012200.RAA27986-at-umic.sunysb.edu} Comments: Authenticated sender is {greg-at-mail.umic.sunysb.edu}
Hi all, I am looking for a special centrifuge tube that allows you to place a TEM grid in it. When the tube is spun the sample settles on the grid. Does anyone know who carries this tube.--Thanks in advance Gregory Rudomen Greg-at-UMIC.SUNYSB.EDU 516-444-3126 University Microscopy Imaging Center S.U.N.Y. Stony Brook
} Once again, I have a bit of a puzzle. I'd like to know } _specifically_ the mechanism that generates continuum X-rays or } "Bremstralung". There is variation among my text references as to the } precise nature of the event that produces this signal. Several texts state } that the interaction is inelastic and results from the slowing of electrons } as they pass near the nucleus (electron-nucleus interaction), while others } state that it is, again, inelastic, and is an primary electron-outer shell } electron interaction (specific references are not given to avoid } finger-pointing). My understanding of the term "inelastic" is that it } refers to electron scattering of primary (beam), backscattered, and } secondary electrons, and not to electron-atomic nucleus interaction. If } someone could take the time to explain in gory detail, I'd appreciate it } very much.
Dear Dwight, The simple explanation of bremsstrahlung is that accelerated charge produces electromagnetic radiation. The mechanism is that when an electron encounters an electromagnetic field, it is accelerated, thus it can radiate. There is no difference whether the field is due to other electrons, nuclei or "wiggler" magnets. Most bremsstrahlung produced by the interaction of electrons with matter is produced by the interaction with nuclei. The field near a nucleus is greater than that near an elec- tron by a factor of Z. Furthermore, from considerations of momentum and energy conservation, there is a higher probability of the reaction
e + M -} e + M + photon
if M is a large mass. Synchrotron light sources use "wiggler" magnets-- an arrangement of magnets which produce fields directed alternately up and down--to direct fast electrons on a sinuous path with a particular frequency. The electrons are accelerated and radiate photons which are more monochromatic than those produced by interactions with nuclei. As was pointed out, any scattering which results in the kinetic energies of the initial particles being greater than those of the same particles in the final state is inelastic. Photon production, changes in internal energy of the target nuclei and nuclear reactions are all examples of inelastic scattering. Yours, Bill Tivol
Very simply, elastic interactions refer to those where energy is not lost (i.e., transformed) during the process. Inelastic interactions involve a "loss" or transformation of energy. Since a portion of the energy of the electron is lost/converted into a bremstrallung photon, the event is by definition inelastic. Now as to where that interation takes place, I will let someone else comment. I thought it was by close encounters with the nucleus.
At 10:04 AM 4/1/97 -0400, you wrote: My understanding of the term "inelastic" is that it } refers to electron scattering of primary (beam), backscattered, and } secondary electrons, and not to electron-atomic nucleus interaction. If } someone could take the time to explain in gory detail, I'd appreciate it } very much. ---------------------------------------------------- Warren E. Straszheim 270 Metals Development, Ames Lab/ISU, Ames IA, 50011 Phone: 515-294-8187 FAX: 515-294-3091
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RE} } Continuum X-ray generation? 4/1/97
Whenever a charged particle is accelerated, it emits radiation. Bremsstrahlung is the term used for the radiation emitted during atomic collisions, where a charged particle is accelerated by the charges it "sees" in its approach. You can find more than you would ever care to know in Jackson's "Classical Electrodynamics," although a more readable (and actually enjoyable) version of the basic physics (quantum electrodynamics) appears in Richard Feynman's "QED."
--------------------------------------
At 10:04 AM 4/1/97 -0400, Dwight Beebe wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
I don't believe we'll ever know the specific mechanism but the macroscopic model we have is apparently built upon an "apparent" bi-modal distribution of "events" we describe as "elastic" and "inelastic", and the (simple) definitions as I interpret them are: "If energy is transferred, then call it inelastic" ... "If very little energy is tranferred, then call it elastic". Another definition of "inelastic" would claim that characteristic information should be the result of an inelastic event.
(... an example of characteristic info gained would be a characteristic x-ray or a auger electron, whereas a backscattered electron, while informative, is not characteristic and is the result of an elastic event ...)
Since the x-ray contiuum is obviously the result of energy transfer it comes under the inelastic event catagory ... yet while it offers no characteristic information (like BSE, at least macroscopically) it has to be attributed to a "continuum" of energy transfers and while we can't "see" what actually goes on (orbital vs. nuclear), why claim one or the other???
... my $0.02 ...
cheerios, shAf
{\/} /\ {\/} /\ {\/} /\ {\/} cogito, ergo ZOooOM {\/} /\ {\/} /\ {\/} /\ {\/} Michael Shaffer, R.A. - University of Oregon Electron Probe Facility mshaf-at-oregon.uoregon.edu -or- mshaf-at-darkwing.uoregon.edu http://darkwing.uoregon.edu/~mshaf/
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There are very good discussions of "Continum X-ray Production" on p. 117 of the Book 'Scanning Electron Microscopy and X-ray Micro Analysis' by Goldsterin, et. al. 2nd Ed, Plenum Press, 1992 (a copy of which should be in every SEM & EMPA lab); and on p. 157-161 of the book 'Electron Beam X-ray Microanalysis' bu Kurt Heinrich, Van Nostrand Reinhold, 1981. Both sources imply that Bremsstrahlung photons are produce by the deacceleration of beam electrons by an interaction with the nuclear field of the atom. In the classical sense such interactions would be considered to be inelastic since they involve a measureble change in energy of the incident electron.
I believe that backscattered electrons are usually considered to be produced by inelastic interactions between the beam electrons and the positive charge of the atoms' core (i.e. the nuclear charge moderated by the tightly-bound inner-shell electrons). This process is described in some detail, both from the classical and the quantum mechanical point of view, by Reimer in his book 'Scanning Electron Microscopy', Springer Verlag, 1985, p. 57-73. Reimer is a pretty well grounded physicist, and so I would think his opinion would be reliable. Reimer treats inelastic scattering processes in similar detail on pages 73-81. The question of the relative magnitudes of the energy transfer involved in elastic and inelastic scattering processes is also discussed on p. 73.
Wilbur C. Bigelow, Prof. Emeritus Materials Sci. & Engr., University of Michigan Ann Arbor, MI 48109-2136 e-mail: bigelow-at-umich.edu; Fx:313-763-4788; Ph:313-764-3321
Rafal Spirydon wrote: } } I am interested in HREM image simulations of III-V semiconductors and I am } looking for any references where I can find the values of the Debye-Waller } temperature factors for such atoms as P, Ga, In, As etc.
Have a look at:
JS Reid, "Debye-Waller Factors of Zinc-Blende-Structure Materials - A Lattice Dynamical Comparison", Acta Cryst A 39 (1983) 1-13.
This reference has Debye-Waller factors for seventeen materials over the temperature range 1 to 1000 K (where appropriate), including GaP, GaAs, GaSb, InP, InAs, and InSb.
Stephen. ............................................................... : Stephen Anderson Australian Key Centre for : : Microscopy and Microanalysis : : Email stephen-at-emu.usyd.edu.au The University of Sydney : : Telephone (+61)-2-9351 7552 NSW 2006 : : Facsimile (+61)-2-9351 7682 Australia : :.............................................................:
We are examining the bacteria Pseudomonas fluorescens (gram-negative soil bacteria) with an SEM. We usually fix with glut, adhere the bacteria to coverslips, dehydrate, CPD, Au/PD coat, and examine. In this case, however, we are having problems with charging and focusing. I haven't had this problem with other bacterial samples. I am going to try osmicating the bacteria but does anyone have any other suggestions?
Thank you in advance,
Ginger Baker EM Lab Manager Dept. APP 250 Vet Med Oklahoma State University Stillwater, OK 74078 (405) 744-6765 FAX: (405) 744-5275 Email: lizard-at-okway.okstate.edu
Rafal Spirydon wrote: } } I am interested in HREM image simulations of III-V semiconductors and I am } looking for any references where I can find the values of the Debye-Waller } temperature factors for such atoms as P, Ga, In, As etc.
Have a look at:
JS Reid, "Debye-Waller Factors of Zinc-Blende-Structure Materials - A Lattice Dynamical Comparison", Acta Cryst A 39 (1983) 1-13.
This reference has Debye-Waller factors for seventeen materials over the temperature range 1 to 1000 K (where appropriate), including GaP, GaAs, GaSb, InP, InAs, and InSb.
Stephen. ............................................................... : Stephen Anderson Australian Key Centre for : : Microscopy and Microanalysis : : Email stephen-at-emu.usyd.edu.au The University of Sydney : : Telephone (+61)-2-9351 7552 NSW 2006 : : Facsimile (+61)-2-9351 7682 Australia : :.............................................................:
The site features a discussion area, links to ESEM sites and images on the net, meeting information, microscopy job list, and an archive of all ESEM related posts to the microcopy listserver. Check it out and send me any feedback you might have.
Scott
------------------ Scott A. Wight email: swight-at-erols.com Homepage: http://www.geocities.com/CapeCanaveral/3429
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Most likely the gold coating is sufficient on top of the bacteria but the bacteria are little "umbrellas" which prevent the gold from forming a continuos coating, connecting the under side of the bacteria and the substrate. It helps if the specimen is osmicated. Infusion of silver nitrate has been used to make the specimen more conductive. The carbon tabs or carbon coated mica solve part of the problem because the substrate is conductive. With an evaporator one could rotate the specimen and evaporate gold from two sources. One source should be at a very shallow 6-10 degrees to the specimen. When sputter coating try placing the specimen at about 45 degrees, give it a somewhat lighter coating and then lift the other side and apply a second coating. Using this method a better coating can form under the specimen. Sometimes people forget to paint a conducting path when the coverslip overhangs the specimen stub, but that is not so much a technical problem, rather it's a self-inflicted wound. Cheers Jim Darley
ProSciTech Microscopy PLUS PO Box 111, Thuringowa QLD 4817 Australia Phone +61 77 740 370 Fax: +61 77 892 313 Great microscopy catalogue, 300+ Links, MSDS ************************ http://www.proscitech.com.au
} } We are examining the bacteria Pseudomonas fluorescens } (gram-negative soil bacteria) with an SEM. We usually fix with glut, } adhere the bacteria to coverslips, dehydrate, CPD, Au/PD coat, and } examine. In this case, however, we are having problems with charging } and focusing. I haven't had this problem with other bacterial samples. } I am going to try osmicating the bacteria but does anyone have any } other suggestions? } } Thank you in advance, } } Ginger Baker } EM Lab Manager } Dept. APP } 250 Vet Med } Oklahoma State University } Stillwater, OK 74078 } (405) 744-6765 } FAX: (405) 744-5275 } Email: lizard-at-okway.okstate.edu }
It is probably your substrate. Try putting them on a 0.2=B5m nucleopore filter. Background is not as pretty but charging is not as much of a= problem. Another alternative (standard practice around here) is to paint a continuous line of conductive carbon or silver paint from the top of the coverslip around the edge to the stub to provide a better grounding path. Do this in a couple of obscure spots to your existing samples and see if it works.=20 You may have to adhere the bact. to the coverslips better as well. Go to the web address at the end of this message and find the "Tips & Tricks link. In the TEM section are a bunch of links called "Stickey" something or other which may be useful. Good luck
At 04:43 PM 4/1/97 -0600, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America=20
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { GO GATORS Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 352-392-1295 ICBR EM Core Lab fax 352-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/ The home of " Tips & Tricks "
It is probably your substrate. Try putting them on a 0.2=B5m nucleopore filter. Background is not as pretty but charging is not as much of a= problem. Another alternative (standard practice around here) is to paint a continuous line of conductive carbon or silver paint from the top of the coverslip around the edge to the stub to provide a better grounding path. Do this in a couple of obscure spots to your existing samples and see if it works.=20 You may have to adhere the bact. to the coverslips better as well. Go to the web address at the end of this message and find the "Tips & Tricks link. In the TEM section are a bunch of links called "Stickey" something or other which may be useful. Good luck
At 04:43 PM 4/1/97 -0600, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America=20
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { GO GATORS Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 352-392-1295 ICBR EM Core Lab fax 352-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/ The home of " Tips & Tricks "
My manager wishes to understand my work. Is there anyone who offers a short course in the fundamentals and basics of TEM analysis for materials applications. Any help will be greatly appreciated.
Thanks, Kim Christensen
Kim Christensen White Oak Semiconductor 600 East Main Street, Suite 800 Richmond, VA 23219 Ph: 804-698-7307 Fax: 804-698-7316
We have two devices for sedimenting material onto a TEM grid. The first is a Beckman Airfuge with the EM90 rotor. The grid is placed on a 5 mm square of 0.025 um nitrocellulose and covered with a film of parlodian. This sandwich is placed in the rotor w/ the grid facing the sample prior to sedimentation.
The second device makes use of the 3 mm tubes for a Beckman ultracentrifuge. The tubes have round bottoms but we make semi- spherical inserts out of epoxy that fit in the bottom and leave a flat surface for the grid to rest on. The inserts are made by pouring a drop of epoxy in a tube, allowing it to polymerize and cutting it out of the tube. We then sand them smooth to reduce their size slightly so they will easily slide into another tube. The inserts are reusable and sterilizable.
Regards, Joe Neilly Microscopy and Microanalysis Abbott Laboratories North Chicago, IL 60064
Does anyone know of a selective etch that will etch away silicon but not silicon nitride? I'm looking at a multilayer sample in plan view for TEM and hope to back etch the si substrate and use the nitride as a stop layer.
Buy your manager the books "Transmission Electron Microscopy", by David B. Williams and C. Barry Carter, Plenum Press, NY. ISBN 0-306-45324-X. And upon receiving it you will want another copy for yourself as well.
On Wed, 2 Apr 1997, Christensen, Kim wrote:
} Dear colleagues, } } My manager wishes to understand my work. Is there anyone who offers a short } course in the fundamentals and basics of TEM analysis for materials } applications. Any help will be greatly appreciated. } } Thanks, } Kim Christensen } Yves MANIETTE Universitat de Barcelona Serveis Cientifico Tecnics Unitat ESCA TEM Carrer Lluis Sole i Sabaris, 1-3 E-08028 BARCELONA ESPANYA
Centralized microscope facilities have become increasingly concerned with cost recovery. Most of us that oversee such facilities are aware that Federal regulations require equal charges for in-house users. This was the subject of a number of recent inquires addressed to this newsgroup. My question is how classes are handled. Do you charge the University department or unit whose class uses the facility? If so, how do you figure the charge? Please reply to this newsgroup or to my e-mail or FAX.
Thanking you in advance for your time
Lewis Coons, Ph.D., Director Integrated Microscopy Center Life Sciences Bldg. Campus Box 526040 University of Memphis Memphis TN 38152-6040 FAX 901 678 4457j
} We are examining the bacteria Pseudomonas fluorescens } (gram-negative soil bacteria) with an SEM. We usually fix with glut, } adhere the bacteria to coverslips, dehydrate, CPD, Au/PD coat, and } examine. In this case, however, we are having problems with charging } and focusing. I haven't had this problem with other bacterial samples. } I am going to try osmicating the bacteria but does anyone have any } other suggestions? } } Thank you in advance, } } Ginger Baker
Ginger, Two suggestions that I've found helpful, both having to do with the non-conductuve nature of glass coverslips: 1) Connect the top surface of the coverslip to the stub with silver paint and cover as much of the top surface of the coverslip as you can sacrifice with silver paint. Leave only the area(s) bare that need to be left uncovered to examine the bacteria. 2) Don't use glass coverslips. Coat both sides of membrane filters in the sputtercoater (careful venting! they like to fly around), mount the coated filters on stubs using carbon-conductive double-sticky discs (Pella, and maybe others). Dry the bacteria from air, alcohol, or HMDS; if from fluid, the final change in drying fluid with bacteria in suspension is dropped directly onto the filter pieces on the stubs, then allowed to dry. Be sure to use filters that have nice holes punched in them (Nucleopore, Poretics, that kind), *not* torturous-path filters like, say, Millipore, or you'll have a hard time telling the difference between bacteria and filter. After drying, sputter coat as usual. Phil
&&& Illigitimi non carborundum &&&&&&&& Philip Oshel Station A PO Box 5037 Champaign, IL 61825-5037 (217) 355-1143 oshel-at-ux1.cso.uiuc.edu *** looking for a job again ******************
Well folks, I have just finished the cost analysis for the EM service lab as well as 13 other services labs that are part of our Biotechnology Center. I am not sure it would stand up to a federal audit, but I generated enough numbers to keep the local auditors and accountants baffled for quite some time.
Essentially we put together all of the cost incurred in running each lab, including building services, operations and maintenance, equipment depreciation, salaries and fringe benefits, and a portion of the administartive costs of our center office. Our grants office was able to come up with those numbers for each of the rooms we occupy. After I had a total figure for each lab I deducted a certain percentage based on the activities in each lab NOT devoted to providing services for which we recover costs. That would include teaching, methods development, student advising etc. We have a generous suplement to our budget from the university so we make no charge for those activities. It was then this adjusted cost of operation upon which I based "true cost" of each service. This cost was then used for a maximum rate determination that we would charge "in house" users. All of our "in house " charges are well below the "true cost" of the service, so I expect we would not have any trouble defending the charges we make to federal grantees, should the feds ever show up at our door. ANyone interested in how I cost out each service should contact me since it is rather complicated and lengthy for this forum.
Good luck to all of you who have to go thru this exercise. } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } At 09:43 AM 4/2/97 -0600, you wrote:
} Dear Colleagues: } } Centralized microscope facilities have become increasingly concerned with } cost recovery. Most of us that oversee such facilities are aware that } Federal regulations require equal charges for in-house users. This was the } subject of a number of recent inquires addressed to this newsgroup. My } question is how classes are handled. Do you charge the University } department or unit whose class uses the facility? If so, how do you figure } the charge? Please reply to this newsgroup or to my e-mail or FAX. } } Thanking you in advance for your time } } } Lewis Coons, Ph.D., Director } Integrated Microscopy Center } Life Sciences Bldg. } Campus Box 526040 } University of Memphis } Memphis TN 38152-6040 } FAX 901 678 4457j } } } } ******************************************************* G.W. Erdos, Ph.D. Phone: 352-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax: 352-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 http://www.biotech.ufl.edu/~emcl/ Home of the #1 Gators ***** "Many shall run to and fro, and knowledge shall be increased" Daniel 12:4
Hello, I am a student of the engineering school of material EEIGM of Nancy (france) realizing at the present time an engineering training period in the material laboratory of the UPC (University of Barcelona-spain). I am working on asbestos characterization by TEM but I have some problems to obtain good sample preparation using the "METHOD 7402-NIOSH":
Sample : Filter with a cellulose ester membrane fibers of asbestos
Sample preparation : 1-remove a section of the filter 2-affix the filter section to a clean glass slide 3-place the slide in a petri dish wish contains several paper filters soaked with 2 to 3 ml acetone. Cover the dish and wait 2 to 4 min for the sample filter to fuse and clear. 4-transfer the slide to a rotating stage inside the bell jar ofa vacuum evaporator. Evaporate a 1 by 5 mm section of a graphite rod 5-place the filter, carbon side down, on a grid (200mesh) in a acetone satured petri dish during hours to disolve the filtre.
the problem is that the carbon film is torn on the TEM grid. Could you help me to resolve this problem??
Thank you in advance
Laurent STEINMETZ ----------------------------------------------------------------------- | Jose M Manero E-mail: manero-at-cmem.upc.es | | Electronic Microscopy Lab | | Department of Materials Science and Metallurgical Engineering, UPC | -----------------------------------------------------------------------
RE: } } We are examining the bacteria Pseudomonas fluorescens } (gram-negative soil bacteria) with an SEM. We usually fix with glut, } adhere the bacteria to coverslips, dehydrate, CPD, Au/PD coat, and } examine. In this case, however, we are having problems with charging } and focusing. I haven't had this problem with other bacterial samples. } I am going to try osmicating the bacteria but does anyone have any } other suggestions? } } Thank you in advance, } } Ginger Baker } EM Lab Manager } Dept. APP } 250 Vet Med } Oklahoma State University } Stillwater, OK 74078 } (405) 744-6765 } FAX: (405) 744-5275 } Email: lizard-at-okway.okstate.edu
I have found it very useful to "pre-coat" either cover slips or Nuclepore polycarbonate filters with either Au or Au/Pd prior to applying samples. This provides a conducting substrate beneath the sample. In the case of Nuclepore filters I coat both sides with ~15-30 nm of metal in a sputter coater unit. With coverslips I always "paint" a thin band of colloidal Ag connecting the upper surface to the underlying SEM stub.
This technique also works for Low Voltage cryo SEM on uncoated specimens. I mix ~1:10 ratio of colloidal Ag with a cryoglue (TBS, OCT, Tissue Tek...) and store this mixture in a 1ml tuberculin syringe without a needle. I apply a drop or two of this, spread it around and use it to affix pieces of my precoated polycarbonate membranes with fresh cells to a Si chip just prior to plunge freezing a sample.
Additionally, the use of Si chips (Ted Pella) instead of cover slips provides a nice smooth conducting surface that cultured cells can be grown on.
Ed Basgall, PhD Penn State University Surface Chemistry Group ejb11-at-psu.edu Materials Research Institute Building Ph: 814-865-0493 University Park, PA 16802-7003 FAX: 814-863-0618
The Appalachian REgional Microscopy Society announces the
AREMS SPRING 1997 MEETING, Thursday & Friday, April 17 & 18
For registration and information contact: Susan Read, BASF Corporation, Sand Hill Road, Enca, NC 28728 Telephone: (704) 667-6353 Fax: (704) 667-6903 E-mail: reads-at-basf.com
MEETING EVENTS AND PROGRAM FOR THURSDAY 17 APRIL:
Noon to 5:00 p.m.: Registration, Comfort Suites, NC 191 at Biltmore Square Mall
1:00 p.m. to 5:00 p.m.: Workshops at Comfort Suites and BASF
Workshop 1: Tours of the BASF Corp. Fiber Products Research & Development Microscopy Laboratories (each tour limited to 15 participants)
Workshop 2: Scanning Electron Microscopy: Philips XL30/EDAX DX4 Integrated System - Philips Electronic Instruments, SEM Laboratory at BASF Fiber Products R & D (limit: 10 participants; If there is enough interest, another session may be added.)
Workshop 3: Basic PC Image Capture in Light Microscopy - Martin Microscope Company, Comfort Suites
Workshop 4: Internet Workshop - Mike Webber of LEO Electron Microscopy, Comfort Suites
6:00 p.m. to 7:00 p.m.: AREMS Social Hour, Enka Lake Club, the Patio (Weather Permitting), Wine tasting: wines from the Biltmore Estate Winery, Asheville, North Carolina Beer Tasting: beers from Highland Brewery, Asheville, North Carolina Light hors d'oeuvres
7:00 p.m. to 9:00 p.m.: AREMS Banquet & Keynote Address, Enka Lake Club, Buffet dinner of prime rib, grilled chicken, vegetables, tossed salad, fruit, dessert, and beverage. Dinner Music by Harpist Carroll Owenby
Early 19th Century Country Fashions: History and Evaluation of Mast Suit - a joint address by: Kathleen Wilson, Research Associate, East Tennessee State University, and curator of the Kings Mountain Cultural Center, Johnson City, Tennessee Patricia Ewer, Textile Conservator, The Biltmore Estate, Asheville, North Carolina Susan Read, Technologist, BASF Corporation, Fiber Products Research & Development, Enka, North Carolina
MEETING EVENTS AND PROGRAM FOR FRIDAY 18 APRIL:
8:00 a.m. to 9:30 a.m.: Registration & Coffee, Enka Lake Club (coffee, juice, fruit & danish will be served)
8:30 a.m. to 9:00 a.m.: Overview of BASF Fiber Products Research and Development - Otto Ilg, Vice-President Technology, BASF Corporation, Fiber Products Division
9:00 a.m. to 9:30 a.m.: The Funny and the Grim Aspects of Microscopy - Don Felty, MicroSolutions
9:30 a.m. to 10:00 a.m.: Examining DNA with a Fullerene Imaging Agent - Alan Cassell, Graduate Research Assistant, Department of Chemistry and Biochemistry, University of South Carolina
10:00 a.m. to 10:30 a.m.: Security and Antiforgery Features in Currency and Security Documents - Dr. Matt Hoyt, Senior Research Chemist, BASF Corporation, Fiber Products R&D
10:30 a.m. to 11:00 a.m.: Break for Visiting with Exhibitors
11:00 a.m. to 11:45 a.m.: AREMS Business Meeting
11:45 a.m. to 12:15 p.m.: Design and Implementation of a Laboratory-Wide Image Management System Using Microscoft Windows NT and Magneto-Optical Storage Technology - Rick McGill, Microscopy and Morphology Research, Eastman Chemical Company, Kingsport, Tennessee
12:15 p.m. to 1:00 p.m.: Snails, Eggs, and Microscopy: Light, SEM, and TEM - Stan C. Kunigelis, Associate Professor of Zoology, Clinch Valley College of the University of Virginia, Wise, Virginia
1:00 p.m.: Closing Remarks & Lunch, Enka Lake Club (Assorted salads and sandwich fixings)
Remainder of afternoon and evening free for visiting Asheville. Visits to the Biltmore Estate are recommended
Hello, We are attempting to immunolabel a ribo- nucleocapsid. We have been successful employing negative staining. This is a single-stranded RNA molecule complexed to many copies of the same protein. We have an antibody that stains westerns very well at 1:10000. Should we try incubating this complex with the antibody at say 1:100 and then apply to a grid or after. I guess it would be easy to try both, but if anyone has experience with this we would appreciate any suggestions. Cheers, Hank Adams Electron Microscopy Laboratory New Mexico State University Las Cruces, NM 88003
Good suggestion! I bought one myself and it is an excellent resource. I also understand that Ron Anderson will be editing a companion to that book which will be a comprehensive look at TEM Specimen Preparation. I think that will be a "must buy" also.
David Henriks TEL: 800-728-2233 (toll-free in USA) South Bay Technology, Inc. 714-492-2600 1120 Via Callejon FAX: 714-492-1499 San Clemente, CA 92673 USA e-mail: henriks-at-southbaytech.com
} } } } } Please visit us at http://www.southbaytech.com { { { { {
Manufacturers of Precision Sample Preparation Equipment and Supplies for Metallography, Crystallography and Electron Microscopy.
Message text written by Yves Maniette } Kim,
Buy your manager the books "Transmission Electron Microscopy", by David B. Williams and C. Barry Carter, Plenum Press, NY. ISBN 0-306-45324-X. And upon receiving it you will want another copy for yourself as well. {
Hi all, I am looking for a special centrifuge tube that allows you to place a TEM grid in it. When the tube is spun the sample settles on the grid. Does anyone know who carries this tube.--Thanks in advance Gregory Rudomen Greg-at-UMIC.SUNYSB.EDU 516-444-3126 University Microscopy Imaging Center S.U.N.Y. Stony Brook
The only one that I am familar with is the Aifuge centrifuge made by Beckman. This airfuge is designed especially for small volume samples and they have a really nicely designed rotor just for putting a grid in the bottom and pelleting your sample right onto your grid
If you already have the airfuge, the rotor is part #347844 and the price was $2,270.00 (back in 1995). I am not quite sure of the price of the airfuge itself, but be warned, Beckman is not known for being reasonable. I think that the price was somewhere in the neighborhood of $20K.
Good Luck, Peggy Bisher
Margaret E. Bisher
NEC Research Institute 4 Independence Way Princeton, NJ 08540. Tel.: (609) 951-2629 Fax: (609) 951-2496 e-mail: peggy-at-research.nj.nec.com
Dear Laurent, When I have done this, I put the filter square on a carbon-film-coated TEM grid filter side UP. The filter gradually dissolves and lowers the carbon film onto the coated TEM grid. The asbestos fibers are caught between the two carbon films. Also, if you use a polycarbonate Nucleopore-type filter, you can skip the "clearing" step and carbon-coat the filter material directly. However, with these, I find you must dissolve the filter in chloroform for 48 hours. You wrote:
} Hello, } I am a student of the engineering school of material EEIGM of Nancy (france) realizing at the present time an engineering training period in the material laboratory of the UPC (University of Barcelona-spain). } I am working on asbestos characterization by TEM but I have some problems to obtain good sample preparation using the "METHOD 7402-NIOSH": } } Sample : Filter with a cellulose ester membrane } fibers of asbestos } } Sample preparation : } 1-remove a section of the filter } 2-affix the filter section to a clean glass slide } 3-place the slide in a petri dish wish contains several paper filters soaked with 2 to 3 ml acetone. Cover the dish and wait 2 to 4 min for the sample filter to fuse and clear. } 4-transfer the slide to a rotating stage inside the bell jar ofa vacuum evaporator. Evaporate a 1 by 5 mm section of a graphite rod } 5-place the filter, carbon side down, on a grid (200mesh) in a acetone satured petri dish during hours to disolve the filtre. } } } } } the problem is that the carbon film is torn on the TEM grid. } Could you help me to resolve this problem?? } } Thank you in advance } } Laurent STEINMETZ Regards, Mary Mary Mager Electron Microscopist Metals and Materials Eng., UBC 6350 Stores Rd. Vancouver, B.C. V6T 1Z4 CANADA tel:604-822-5648, fax:604-822-3619 e-mail: mager-at-unixg.ubc.ca
H. ADAMS wrote: } } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Hello, We are attempting to immunolabel a ribo- } nucleocapsid. We have been successful employing negative staining. This is } a single-stranded RNA molecule complexed to many copies of the same } protein. We have an antibody that stains westerns very well at 1:10000. } Should we try incubating this complex with the antibody at say 1:100 and } then apply to a grid or after. I guess it would be easy to try both, but } if anyone has experience with this we would appreciate any suggestions. } Cheers, } Hank Adams } Electron Microscopy Laboratory } New Mexico State University } Las Cruces, NM 88003 } } http://www.nmsu.edu/Research/artsci/public_html/eml/ } } 505-6463600
Hi Hank,
If I understand correctly you want to label the nucleoprotein on your RNP complex. When I label the measles nucleoprotein on purified RNPs I do it like this : adsorb the RNPs onto a formvar carbon coated grid, remove the excess and immediately add the first antibody (anti protein) generally diluted 1/100 or more (I do 3 dilutions in fact). wash add the second antibody (coupled to gold) wash negative stain as usual
Normally if the first antibody works well, you don't need the second one, at the EM level you can see the RNP much bigger because of the IgG fixed on it. (after staining of course)
Good Luck Daniele SPEHNER Electron Microscopy Laboratory Etablissement de Transfusion Sanguine 67065 Strasbourg Cedex - FRANCE
Wyeth-Ayerst Research, a major division of Fortune 100 American Home Products Corporation, has an opportunity at our pharmaceutical research facility in Chazy, New York for a Scientist in our Imaging Laboratory.
The selected candidate will be responsible for operating a transmission electron microscope (TEM), scanning electron microscope (SEM), and processing images. Resposibilities include processing, embedding, and sectioning tissues in support of electron microscopy services, as well as performing histological techniques. The incumbent must keep records in compliance with SOP's and GLP's and maintain laboratory instruments in accordance with SOP's, troubleshooting equipment problems as needed.
Requirements include a B.S./M.S. with 2-7 years relevant experience and familiarity with all techniques necessary for the operation of TEM, including tissue preparation and image processing. SEM experience is desireable.
The Wyeth-Ayerst Drug Safety Division is located between the foothills of the Adirondack Mountains and the shores of Lake Champlain. The area offers a wide range of outdoor recreational activities while being a short distance from Lake Placid, NY; Burlington, VT; Montreal, Quebec, and Plattsburgh, NY. Wyeth-Ayerst Research offers an excellent compensation and benefits package in a highly professional environment. Please send your resume with salary requirements to:
Mr. Lou Ballester Human Resources Wyeth-Ayerst Research 641 Ridge Road Chazy, NY 12921
You wrote: } } Hi all, } I am looking for a special centrifuge tube that allows you } to place a TEM grid in it. When the tube is spun the ......
Hi Greg (and all),
In the last Fullam catalog I have (1992-93), EFFA Centrifuge Tubes are pictured on page 49. Cost of the tubes then was $135.00/balanced pair (#11450). They also make some to hold No. 00 BEEM capsules, which are the ones I've used in the past (pretty nifty). These are good up to 6000g. They also listed some others that are good up to 34,000g with which I've had no experience. Don't know if they're still available, you'll have to check.
Heather Owen
p.s. I have no connection with Ernest F. Fullam, Inc. - just a satisfied customer.}
Heather A. Owen, Director Electron Microscope Laboratory Department of Biological Sciences University of Wisconsin - Milwaukee (414)229-6816
To all of you who have asked for details on our cost accounting:
More of you responded that I thought might so I have posted the data at our WWW site. I didn't know how to email spreadsheets.
Some places my columns are a little screwed up but I think you can figure it out.
so go to http://www.biotech.ufl.edu/~emcl/cost.html
Let me know if you do not have access to the WWW and I will fax. ******************************************************* G.W. Erdos, Ph.D. Phone: 352-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax: 352-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 http://www.biotech.ufl.edu/~emcl/ Home of the #1 Gators ***** "Many shall run to and fro, and knowledge shall be increased" Daniel 12:4
Dear Microscopy Folks: I am Candace Haigler, Director of the EM Lab at Texas Tech University, writing this message together with Mark Grimson, EM Technician, who is a member of your Internet group. We find ourselves facing a potential problem about which we need your advice.
Our EM lab is now fairly stable, being purpose-built underground as an attachment to the main Biology building (it sticks out beyond the above-ground footprint of the building). Our only current problem is that our roof is a brick patio that attracts skate-boarders. When they are skate-boarding, we cannot take pictures due to vibrations, but this is a sporadic problem and we can chase them away.
Now we find that the university is making a Master Plan, the draft of which shows a major service road to a new 1000 space parking deck coming very close to our building. We estimate that the road will be within 50 feet, or even closer, to the below-ground EM lab. Given our existing problem with skate boarders, we are very worried that this road will essentially destroy the utility of the lab.
We solicit your help in: (1) Sharing knowledge about similar situations (2) Pointing us to the best published sources about EM lab design, particularly in regard to vibration and preferred distance from nearby roads (3) Pointing us to any expert EM lab design firms from whom we might get information
We are very concerned about this situation, and will greatly appreciate your help. You may reply to Mark at the address shown above, or my personal e-mail address is brchh-at-ttu.edu.
Sincerely, Candace Haigler Professor and Director of the Electron Microscopy Laboratory
We have recently purchased a Polaron CPD and wanted to hear from anyone else who has one. I had used one at a University, at which time once I loaded the specimen chamber and shut the back, opened the fill valve, the CO2 leaked out the front window. I was concerned and drained out the CO2. I went to find the instructor, and asked why this was happening. I assumed a seal was faulty, but she told me that the boat wasn't loaded properly. The back had closed nicely, and I really didn't think that it was not loaded properly. Once the chamber was reloaded, the CO2 tank was empty and I could not finish the run. Once the CO2 tank was replaced weeks later, I received a phone call saying that the seal was damaged and that I would have to wait before the new ones came in. To make a long story short, we now have a Polaron CPD and I am having the same problem. Sometimes when I load it, it leaks out the front. Everything seems to be fitting properly, but on occasion, upon filling, it leaks out the front window. The only reason that I am posting this question to the listserver and not to the manufacturer, is because another user thinks that this is normal?! Even if the boat was not fitting properly, should the vessel still leak out the front? I would appreciate any enlightment on this problem. It is a brandnew CPD and thus I have my doubts that the seal would be damaged already. Perhaps it is just not seated properly.
P.S. If anyone has recently purchased a Polaron CPD and finds out that the seal inside the chamber door keeps falling out, a piece of teflon tape around the seal works wonders!
Susan
Susan Carbyn Atlantic Food and Horticulture Research Centre Agriculture and Agri-Food Canada Kentville, Nova Scotia B4N 1J5 Canada
Can anyone please comment on the storage properties of common EM/LM fixatives. We have some older 1% glut/4% formaldehyde in PO4 buffer. Is there some rule of thumb that people are using, should we analyze before use, are there some references we can access? Thank you in advance for your comments.
} Now we find that the university is making a Master Plan, the draft of which } shows a major service road to a new 1000 space parking deck coming very } close to our building. We estimate that the road will be within 50 feet, } or even closer, to the below-ground EM lab. Given our existing problem } with skate boarders, we are very worried that this road will essentially } destroy the utility of the lab. } Dear Candice et al., This sounds like a real disaster in the making. Our facility is several hundred meters from some major roads, and we were quite worried about vibrations from truck traffic. The effects of traffic depend crit- ically on the nature of the soil between you and the road. Bedrock will transmit vibrations very well; whereas damp clay will absorb much of the energy. The good news is that there may not be much traffic except for a few times during the day, and that you may be able to convince the university to put some vibration-damping material at the bottom of the roadbed. I don't know what is available, but maybe a layer of poly- urethane (which is a good vibration absorber) could be cost-effective solution--especially if there is a source of recycled plastic locally. Good luck. Yours, Bill Tivol
As a result of my comments on bremsstrahlung, inelastic, and elastic scattering, questions have been raised concerning energy loss as an electron's path is changed. That is,it gets swung around the nucleus somewhat like a comet gets swung around the sun, and since this involves a change in direction, which in turn amounts to deacceleration, should involve some loss in energy. As I hope I implied in my original comments, I do not consider myslef to be an authority on matters of electron-atom interactions. All I intended to do was to give a couple, what I considered to be clear and useful, references on the subject. And so, in an attempt to answer the more recent question I quote from Reimer, p. 73 of his book 'Scasnning Electron Microscopy':
"During elastic scattering of electrons, as discussed in Sect. 3.1, the sums of momenta and of kinetic energy of the collision partners are conserved. The energy loss of the incident electrons and the kinetic energy transferred to the nucleus can be neglected for the electron energies used in SEM since the electron mass is so much smaller than thet of the nucleus. Even when 30 keV electrons are scattered through an angle of 180=B0, the energy transferred to a Cu nucleus is only of the order of one electronvolt, and such scattering processes have a much lower probability than inelastic processes with energy losses larger than 5 eV. Only for electrons in the MeV region can the energy transferred bvecome larger than the displacement energy necessary to dislodge an atom from its lattice site into an interstitial position, which is of the order of 10-30 eV."
I hope this will help clarify the matter. For more details refer to Reimer's book.
Wilbur C. Bigelow, Prof. Emeritus Materials Sci. & Engr., University of Michigan Ann Arbor, MI 48109-2136 e-mail: bigelow-at-umich.edu; Fx:313-763-4788; Ph:313-764-3321
Candice/all: We here at Dow are in a well-isolated facility which is a result of demonstrating that passing trucks would give problems to our NMR spectrometers and microscopes. Without going into the full explanation, the argument was made based on empirical data: We had large trucks rumble by our existing facilities during data acquisition and compared the results to the same experiment run during a known quiet time. The loss of information was documented and recast in terms of monetary cost for reduced data quality. In our case, the financial penalty of reduced resolution/sensitivity justified the extra cost of closing a major local thoroughfare.
My suggestion would be to get someone from a trucking company to come by and drive their truck in the approximate location of the service drive to document the problems, then see if the U. can come up with an alternate access route to the ramp (moving the entire ramp would be better, but probably less likely). The fact that you have a few hundred thousand dollars tied up in sensitive equipment suggests that the U. recognizes the value of your work and would hopefully be willing to accommodate the situation. Consider the bad press they would get for compromising their research reputation in the name of a car park!
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } Hi Susan, Try filling the chamber very slowly. I have the same problem on my BioRad CPD. Changing the seals didn't help.
} Hi there, } } We have recently purchased a Polaron CPD and wanted to hear from } anyone else who has one. I had used one at a University, at which time } once I loaded the specimen chamber and shut the back, opened the fill } valve, the CO2 leaked out the front window. I was concerned and drained } out the CO2. I went to find the instructor, and asked why this was } happening. I assumed a seal was faulty, but she told me that the boat } wasn't loaded properly. The back had closed nicely, and I really didn't } think that it was not loaded properly. Once the chamber was reloaded, } the CO2 tank was empty and I could not finish the run. Once the CO2 } tank was replaced weeks later, I received a phone call saying that the seal } was damaged and that I would have to wait before the new ones came in. } To make a long story short, we now have a Polaron CPD and I am having } the same problem. Sometimes when I load it, it leaks out the front. } Everything seems to be fitting properly, but on occasion, upon filling, it } leaks out the front window. The only reason that I am posting this } question to the listserver and not to the manufacturer, is because another } user thinks that this is normal?! Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already. Perhaps it is just not } seated properly. } } } } Gregory Rudomen Greg-at-UMIC.SUNYSB.EDU 516-444-3126 University Microscopy Imaging Center S.U.N.Y. Stony Brook
Contact Dr. Judy Murphy (expert in design of EM labs). She may be able to guide you. 209 474-5284
Also check Chapter 1, Setting Up An Electron Microscope Facility in Procedures in Electron Microscopy, AW Robards and AJ Wilson, eds, John Wiley & Sons, New York. While it doesn't address skateboarders and roads per se, it may give you ammunition to fight the administration.
Sara E. Miller, Ph. D. P. O. Box 3020 Duke University Medical Center Durham, NC 27710 Ph: 919 684-3452 FAX: 919 684-8735
I am currently working on gold labelling project involving monolayr. I use LR GOLD and LOWICRYL and do my embedding and polymerization in the UVC 2 cryochamber by TED PELLA. My problem is that the blocks polymerize with in 1 hr. Has anyone experienced this? What can I do about it. I want slow polymerization.
I follow the protcols provided and at minus 10 C.
******************************************************** * Raj Patel * * Dept. of Pathology * * Robert Wood Johnson Medical School * * 675 Hoes Lane, Piscataway, NJ 08854 * * * * voice (908) 235-4648; Fax -4825 * * E-Mail rpatel-at-umdnj.edu * ********************************************************
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ed
} } Hi there, } } We have recently purchased a Polaron CPD and wanted to hear from } anyone else who has one. I had used one at a University, at which time } once I loaded the specimen chamber and shut the back, opened the fill } valve, the CO2 leaked out the front window. I was concerned and drained } out the CO2. I went to find the instructor, and asked why this was } happening. I assumed a seal was faulty, but she told me that the boat } wasn't loaded properly. The back had closed nicely, and I really didn't } think that it was not loaded properly. Once the chamber was reloaded, } the CO2 tank was empty and I could not finish the run. Once the CO2 } tank was replaced weeks later, I received a phone call saying that the seal } was damaged and that I would have to wait before the new ones came in. } To make a long story short, we now have a Polaron CPD and I am having } the same problem. Sometimes when I load it, it leaks out the front. } Everything seems to be fitting properly, but on occasion, upon filling, it } leaks out the front window. The only reason that I am posting this } question to the listserver and not to the manufacturer, is because another } user thinks that this is normal?! Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already. Perhaps it is just not } seated properly. } } P.S. If anyone has recently purchased a Polaron CPD and finds out that } the seal inside the chamber door keeps falling out, a piece of teflon tape } around the seal works wonders! } } } Susan } } } Susan Carbyn } Atlantic Food and Horticulture Research Centre } Agriculture and Agri-Food Canada } Kentville, Nova Scotia B4N 1J5 } Canada } } Phone: (902) 679-5566 } Fax: (902) 679-2311 } E-mail: carbyns-at-em.agr.ca
Ed Basgall, PhD Penn State University Surface Chemistry Group ejb11-at-psu.edu Materials Research Institute Building Ph: 814-865-0493 University Park, PA 16802-7003 FAX: 814-863-0618
Job description: SEM operation and sample preparation in support of DRAM fabrication and failure analysis. The candidate will perform: 1. Delayering/deprocessing by RIE/Plasma, wet etch and parallel polishing/face-lapping. The deprocessing will require the use of chemicals (acids and solvents) within a wet lab. 2. Optical microscope inspections by bright field and differential interface contrast. 3. Cross section preparation using mechanical polishing, fracturing, and focused ion beam (FIB) techniques. 4. SEM inspections by secondary electron imaging, back-scatter imaging and energy dispersive spectroscopy (EDS). Education and experience: The candidate should ideally have an associates degree and several years of semiconductor SEM experience. If there is no semiconductor SEM experience, the candidate should possess a college degree in a microscopy related field (i.e. biology/geology/materials science/etc) and be actively using SEM and optical microscopy techniques. Good eye-hand coordination, communication skills, and attention to details are required. Candidates should be highly motivated, self directed, interested in learning new skills an effective working alone or in a team environment. ******************************************* Bart Seefeldt White Oak Semiconductor 600 East Main Street, Suite 800 Richmond, VA 23219 Ph: 804-698-7225 Fax: 804-698-7316 E-mail: seefeldb-at-whiteoaksemi.com *******************************************
Has anyone had experience using a Pixera camera on a TEM? I am attempting to put one on a Zeiss 902 with a C mount connection. If the camera is directly mounted, the focal length is not correct. Does anyone know of an adapter than can be used so theimage can be focused correctly?
Thanks for any suggestions.
Nancy R. Smith Microscope and Graphic Imaging Center California State University, Hayward
} } We have recently purchased a Polaron CPD and wanted to hear from } anyone else who has one. I had used one at a University, at which time } once I loaded the specimen chamber and shut the back, opened the fill } valve, the CO2 leaked out the front window. I was concerned and drained } out the CO2. I went to find the instructor, and asked why this was } happening. I assumed a seal was faulty, but she told me that the boat } wasn't loaded properly. The back had closed nicely, and I really didn't } think that it was not loaded properly. Once the chamber was reloaded, } the CO2 tank was empty and I could not finish the run. Once the CO2 } tank was replaced weeks later, I received a phone call saying that the seal } was damaged and that I would have to wait before the new ones came in. } To make a long story short, we now have a Polaron CPD and I am having } the same problem. Sometimes when I load it, it leaks out the front. } Everything seems to be fitting properly, but on occasion, upon filling, it } leaks out the front window. The only reason that I am posting this } question to the listserver and not to the manufacturer, is because another } user thinks that this is normal?!
This is EXACTLY THE SORT OF MATTER TO PUT ON THE LISTSERVER.
Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already.
We have a Polaron CPD 3000 and the seals often leak, particularly the one on the window. I have taken the CPD apart myself to check it. The window seal leaks even though there are no physical defects on it. And you can replace the same seal in the window and next time it will not leak, indicating no permanent damage. So it is not damaged in the sense that a badly loaded boat has pushed up against it and nicked it so it leaks. In fact I can't see how loading the boat would ever make a difference to the integrity of either of the seals, considering how they are located completely inside grooves.
BUT the door seal can be physically damaged by hard particles (say bits of cover slip) getting washed out of the chamber and locating at the sealing surface so they nick the seal as you screw up the door. SO you need to go carefully around the DOOR sealing surface and screw thread with say ethanol on a cotton bud once a week.
AND the seals on the drain valve on the bottom need regular cleaning for the same reason. Grit washes down the drain and scores the sealing surface and O rings in the valve. In fact if you have never done it, go and clean the door seal and drain valve right away. You'll be surprised at the gunge that will be there.
My explanation is to do with seal elasticity. When the CPD is cooled before being pressurised, the neoprene seals lose much of their elasticity and do not expand properly to seal when the chamber is pressurised. So the solvent leaks out and as the seal stays cold it will never seal properly as of course you keep the chamber cold to ensure fast fluid transfer.
The remedy according to Me is to pressurise the CPD BEFORE COOLING IT. BTW we heat and cool the CPD by circulating water through the jacket using a waterbath heater/mixer (Lauda type T, -20deg to 100 deg C.) which has a small centrifugal pump on it and which sits in a 2 Litre stainless tank. At the start we fill up the small tank with ice and add enough water to cover the heater/pump on the mixer. Then we start the pump with the heater off and chill the CPD that way. When we want to warm the CPD we toss away the ice water, replace it with tap water -at- 20 deg or so and turn on the heater which warms the waterbath towards 50 deg.
Notwithstanding all of the above, since the CPD only works if all the seals are good, you must keep a couple of seal kits in your lab ready for quick repairs. Failure of the seals is usually discovered after some poor soul has spent weeks on their experiment and days on processing their tissue only to find the CPD leaks. If you have the capacity to quickly replace any of the seals so their experiment survives your reputation will be much enhanced!
Mel Dickson E.M. Unit, University of NSW Sydney, Australia
} We solicit your help in: } (1) Sharing knowledge about similar situations
I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off.
The problem will vary according to the soil type between your lab and the new road. Ideally the road would be on swampy ground and your lab on a platfrom carved out of granite. That would give good decoupling. But basically, any effective solution in you laboratory will cost several thousands per instrument, as each instrument will need to be relocated on some heavy support (thick steel plate, thicker concrete slab, mass is what you need) with some very flexible mounts under it (air-springs are ideal). It may be more cost effective to route the road further away.
} (2) Pointing us to the best published sources about EM lab design, } particularly in regard to vibration and preferred distance from nearby roads
} (3) Pointing us to any expert EM lab design firms from whom we } might get information
The classic reference work is "Design of the Electron Microscope Laboratory" by Ronald H Alderson 1975. It is Volume 4 in "Practical Methods in Electron Microscopy" Editor Audrey M Glauert. American Elsevier ISBN 0444 1087 6. Was still available two years back whem I bought my second copy to share with the architect for my new laboratory. Pages 68-86 deal with mechanical vibration and decoupling/damping systems } } Mel Dickson } } } }
} } Can anyone please comment on the storage properties of common EM/LM fixatives. } We have some older 1% glut/4% formaldehyde in PO4 buffer. Is there some rule } of thumb that people are using, should we analyze before use, are there some } references we can access? } Thank you in advance for your comments. } } Regards, } Ken Baker } } The aldehydes oxidise to acids - formic or glutaric. The reaction is less at lower pH so is accelerated when you buffer the solution to pH7. Our rule is to buffer the fixative just before we use it and discard any buffered fixative older than a week.
Thanks to all those who responded in tracing Edington's Practical electron microscopy book. Tech Books distributes it through their distributor "Ceramic Book and literature Service (CBLS)."
A Lab6 cathode running on a Hitachi H7100 TEM (manufacturers brand) developed a bright rectanglar-shaped emmision pattern over the last 50-100 hours of operation, although it can be biased back to a small circular spot. We hadnt seen this before. The tip of the cathode shows a distinctly bar-shaped tip when viewed in a light microscope.
The cathode has been very stable over its 450 hours of life, although it seems to move a little vertically as it heats up. There is plenty of Lab6 left.
Does anyone know what causes the pattern, and what are the chances of getting back to a squarish/maltese cross type image if we, say, run it a bit hot for a while?
regards, Sally ---------------------------------------------------------------------- Sally Stowe |Email: stowe-at-rsbs.anu.edu.au Facility Coordinator |Post: ANU Electron Microscopy Unit |ANUEMU (RSBS) Ph 61 6 249 2743 |Australian National Univ. FAX 61 6 249 4891 |Canberra, http://online.anu.edu.au/EMU/home.htm |AUSTRALIA 0200
... snips } user thinks that this is normal?! Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already. Perhaps it is just not } seated properly. } } P.S. If anyone has recently purchased a Polaron CPD and finds out that } the seal inside the chamber door keeps falling out, a piece of teflon tape } around the seal works wonders! } } } Susan } } } Susan Carbyn } Atlantic Food and Horticulture Research Centre } Agriculture and Agri-Food Canada } Kentville, Nova Scotia B4N 1J5 } Canada } } Phone: (902) 679-5566 } Fax: (902) 679-2311 } E-mail: carbyns-at-em.agr.ca
Well, I wouldn't consider leaking from the from window normal - get the supplier/manufacturer to sort it out.
However, it might be a handling problem rather than an equipment fault. Make sure you go through all the steps slowly, as sudden temperature changes in particular could be causing differential expansion.
I'd also be unhappy about the tape on the door seal. Although unlikely to cause problems, there is a remote chance it might. A better solution is to hold the metal seal edge on and give it a tap against a hard surface - the idea is to make it slightly oval, so it grips its seating.
Our Polaron E3000 CPD was bought in 1976 and is still going strong, wuite a lot of use too!
When you say Teflon tape around the seal, I in=magine you mean peripherally rather than through the hole in the middle?!
My experience is that the Doughty/Dowty? seal, the main metal ring plus nitrile (?), tends to split regularly with acetone, every 2-4 months or so. If everything has worked once, then assembly should be ok, it is possible for things to loosen but in my experience that is very unusual. You need to look at the inner face of the seal for any imperfection. I don't know if you get any specail tool, but I made up a steel oblong gizmo which is like a big screwdriver blade whichand gets turned gently with an adjustable wrench.
Hey! I've just realiased, we should have another party for its 21st birthday!!! We get a lot of parties around here, folks!
Many years ago JEOL News published an article on the design of the EM rooms at the John Innes Institute in the UK. As far as I can recall this dealt in some detail with vibration transmission. We based the design of our EM rooms on this and we have had no vibration problems.
Robin H Cross Director : EM Unit, Rhodes University, Grahamstown, South Africa eurc-at-giraffe.ru.ac.za - tel: +27 461 318168 - fax: +27 461 24377
Hi Candace. There was a discussion a while back on this topic which has been archived at the "Tips & Tricks" site. Go to the web address listed at the end of this message and follow the "Tips & Tricks link. Proceed to the Miscl. link and there it is at the top of the list. Good luck.
At 10:07 AM 4/3/97 +0131, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { GO GATORS Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 352-392-1295 ICBR EM Core Lab fax 352-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/ The home of " Tips & Tricks "
Looking for experienced TEM Service Engineer to service Philips 420 in Kansas City Area. Contact Pete Dondl at sylviapns-at-worldnet.att.net P & S Products, Inc.
In reply to: ================================================== I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off. ================================================== There have been instances where the problem with the rail line has been less due to ground vibrations than to the creation of a transient magnetic field problem that correlated with the passage of a train. In about 1969, there was an SEM installed at a Philadelphia university near the main passenger line of AMRAK and Conrail, and the real problem was more related to the magnetic field (trains were pantograph (electrically) powered) than to vibrations. The problem was ultimately "solved" only by moving the microscope. So don't forget the magnetic field problem potential as well.
Chuck
=================================================== Charles A. Garber, Ph. D. Ph: 1-(610)-436-5400 President 1-(800)-2424-SPI SPI SUPPLIES FAX: 1-(610)-436-5755 PO BOX 656 e-mail: cgarber-at-2spi.com West Chester, PA 19381-0656 USA Cust. Service: spi2spi-at-2spi.com
Look for us! ############################ WWW: http://www.2spi.com ############################ ==================================================
Larry's comment about knocking the seal out of 'round' reminded me - I forgot to mention in my earlier message that I do this in a large workshop vise. A gentle squeeze works a treat! (and that is not a naughty comment!). Plus, the squeeze is more controllable, if at first it doesn't work, you can go back for more.
We have a Phillips 300 TEM available to anyone who wants it. It was purchased in 1980 and completely renovated in 1994. It comes with cooling system and a standard diffusion pump instead of the original mmercury pump. It does need work on the condenser lens electronics, but otherwise works beautifully. It is free, anyone interested will just need to come and get it as we need it out asap.
Thanks!!
Cheri Owen Detroit Neurotrauma Institute Wayne State University Detroit, Mi 313-577-4648
Due to the overwhelming number of personal responses I received on my CPD question, I thought I would summarize some of what I found out.
My first reaction to all the responses, is that the Polaron CPD E3000 is used by many people in the field of EM and thus must be an excellent product.
Many responses indicated that for years, people have had no problems with their CPD's leaking. It hasn't been until fairly recently, after years of usage, that people have experienced their "Downty" seals (The seal in the window), leaking. I would imagine that after years of usage, they would need to be replaced, but I was shocked at the number of people who seem to be replacing them fairly regularly. One person stated that after several good runs (varies from 5 to 10 or more), they get a leak. Personally, I don't think 5 to 10 runs is a lot! This would suggest to me, that perhaps the seals are not as "good" quality as they used to make them?
Some suggested that temperature changes result in contraction/expansion of the seals which might explain the leaking. Another response indicated that the seals will dry up and crack easily. Since our machine and all of it's parts are brand new, the age of the seals are not a factor.
To test for leaks, a lot of people recommended pressurizing the vessel every time, prior to loading it with samples. This however, does not necessarily guarantee that when you load the chamber with your samples that it will NOT leak. One person felt that the problem was a handling one rather than an equipment fault. This would probably relate back to the temperature changes by filling a chamber too quickly that would result in differential expanding of the window Downty seal.
Many people replied to me and said that the Teflon tape around our door seal may be the problem or may cause future problems. No one said exactly why this was, other than a prompt reply I got from the manufacturer. The manufacturer thought that the tape may break off and become lodged in the drain valve. We only used a tiny piece wrapped tightly around the outside of the ring (Not around the entire circumference of the seal as some misinterpreted), but rather around the outer edge - like a piece of tape or bandaid one might wrap around a ring to make it fit their finger.
Many people have bent their door seal so that it's oval shape would hold it in place.
I appreciate all of you who responded and especially to the manufacturer! I didn't contact the manufacturer first thing, because I wanted to see if there were other people with some similar experiences that could help me "quickly" solve the problem.
The manufacturer assured me that if the boat wasn't loaded correctly, the door would not close at all. The most likely cause of the problem is from the Downty seal which is not manufactured by them, but rather purchased in. They have had batches know to be faulty and in such instances, they would discard and return to the original manufacturer. They state: if the front window leaks, there are only 2 possibilities:
1. The front is not screwed tight enough 2. The Downty seal is faulty
Finally, the manufacturer said that recently they had a batch of Downty seals which were of the wrong material and very quickly deteriorated with the dehydration solvents being used. As for the door seal needing to be bent, the business manager has requested the design team to review the way the seals are held in place and that some good news may come from the problems others have shared on this topic, on this list server!
As one colleague from the list server wrote: "Amazing the number of responses with the same problem. And we toil away thinking we're the only ones with the weird difficulties".
Thanks again to all who replied. Although my problem had an easy solution, it has given me insight on lots of other situations!
Susan
Susan Carbyn Atlantic Food and Horticulture Research Station Agriculture and Agri-Food Canada Kentville, Nova Scotia B4N 3R2 Canada
This last piece of information was passed along to me by the Manufacturer of the Polaron CPD.
Please ammend the e-mail or notify the customer with the faulty CPD that the seals used were not in fact made by DOWTY but of similar design, from another manufacturer, these in fact were of the wrong material so in practice the material was not faulty but the manufactured item itself was below spec.
Susan
Susan Carbyn Atlantic Food and Horticulture Research Station Agriculture and Agri-Food Canada Kentville, Nova Scotia B4N 1J5 Canada
We are interested in obtaining a carbon coated grid but in a "butterfly" or folding configuration instead of a single grid. We have some powders that are radioactive and though our microscope (TEM) is already contaminated we would like to reduce the potential for further contamination. I was thinking that a butterfly grid with carbon on both sides may help.
Any thoughts? Does anyone sell such a beast? Any other ideas for reducing the amount of particles that may come off?
Most of my work is with metal samples and I haven't worked with carbon coated grids much. Can we make something like that easily? I saw the discussions about holey carbon films but that is not quite what we are after!
TIA
John Vetrano Pacific Northwest National Laboratory Richland, WA 99352 js_vetrano-at-pnl.gov
We are in the process of purchasing a TEM for biological application. The models we are planning to check on are Hitachi H-7500, JEOL JEM-1220, and Philips CM100 BioTWIN. We would like to hear the opinions of people who have experience with these models. What are the pros and cons? Thank you very much.
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Dear Angus:
I posed your question to Bernie Kestel from Argonne National Laboratory and he offered the following:
" RE} selective etch for silicon 4/4/97
I once used the following solution on Si with a silicon oxide layer. Only the Si was polished away on the South Bay Technology 550 B jet polisher. 60 ml. HF, 90 ml. sulphuric acid, 100 ml. butyl cellosolve (2-butoxyethanol), 500 ml. methanol. Conditions: 80 volts, 50 ma., -45 degrees Centigrade, (dry ice/methanol), slightly "slow" pump speed ~ 4. Used green LED light source. 150 micron test hole to set auto trip at 6.5 on front panel by adjusting detector bias (rear knob), to get those conditions. If silicon nitride is conductive, this may not work."
You can get additional information on the Jet Polisher (Now Model 550D) on our web site at http://www.southbaytech.com.
DISCLAIMER: As we do manufacture the Model 550D Single Vertical Electropolisher, I obviously have a vested interest in promoting its use.
David Henriks TEL: 800-728-2233 (toll-free in USA) South Bay Technology, Inc. 714-492-2600 1120 Via Callejon FAX: 714-492-1499 San Clemente, CA 92673 USA e-mail: henriks-at-southbaytech.com
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To any who can help...... I need to find a way to project some 4x5 negatives onto a screen for quantitation purposes. I know that such projectors used to be around, but here at Mayo they are history. I need the projector because I have many low mag micrographs that I am quantitating, and making larger (16x20) prints is not really feasable. Any leads as to where I can borrow, buy or make such a projector would be very helpful!!!
TIA Eugene Krueger GI Research Mayo Foundation krueger.eugene-at-mayo.edu
} We are interested in obtaining a carbon coated grid but in a "butterfly" or } folding configuration instead of a single grid. We have some powders that are } radioactive and though our microscope (TEM) is already contaminated we would } like to reduce the potential for further contamination. I was thinking that a } butterfly grid with carbon on both sides may help. } } Any thoughts? Does anyone sell such a beast?
I haven't seen this advertised, but one of the suppliers on our list will certainly correct me if they sell these.
} Any other ideas for reducing the } amount of particles that may come off? } A formvar or collodion film could work--especially if charging is not a problem. You don't say whether the sample is conductive or not.
} Most of my work is with metal samples and I haven't worked with carbon coated } grids much. Can we make something like that easily?
Yes, [even I can make one :-)]. Two procedures are possible: 1) Cleave a mica sheet to expose a fresh surface, evaporate carbon onto that surface, lower the carbon-coated mica at an angle into a staining dish filled with distilled water to float the film off the mica
Prepare the grids by rinsing in dilute nitric acid then distilled water, place open folding grids, inside down, onto the floating carbon film (care- fully), place a square of filter paper over the grids & film and lift off the surface of the water (again, carefully). 2) Buy or make a solution of formvar in ethylene dichloride--the proper dilution will depend on the final thickness of the film. Take a clean glass microscope slide and apply a very light coating of finger or nose grease. Dip the slide into the solution, let the excess drip off and let the solvent evaporate in dry air--if you are in a humid environment, you will have to fill a volume with dry N2. When the film has hardened, score the slide with a razor blade very near the edges to give a film which is attached only to one surface of the slide. Lower the slide at an angle into 60 deg C water to float the film off, prepare, place & pick up grids as above. Evaporate carbon onto the formvar-coated grids. The first method gives a thinner film, but it may not prevent the escape of small bits of powder; the second method gives a less porous film. When you want to use the grids, you will have to cut them out to remove them from the filter paper. Be careful to make sure that the film adheres to the whole surface of the grid during this and the folding process. Good luck. Yours, Bill Tivol
Dear Colleagues: Can any of you recommend a good reference book on preparations of macromolecules, e.g.,DNA and proteins, for electron microscopy? I am looking for a comprehensive review book which describes methods of positive, negative staining, rotary shadowing of biological macromolecules. Thank you in advance.
If I understand your question correctly, I think that you have several options.
1) You could make your own carbon coated folding grids using the same procedures used for single grids ( check for example "Techniques for Electron Microscopy" Ed. Desmond Kay ). The techniques (there are a number of them) are relatively simple.
2) Could you use a single carbon coated grid and then deposit carbon (by vacuum evaporation) on top of your particles on the grid ?. This would serve the same purpose as the carbon coated folding grids. You might end up contaminating your evaporator however.
No matter what you do, you will still run the risk of contaminating your scope since some of the film might break during observation. Also , keep in mind that the increased thickness (two carbon layers), will decrease your resolution. This might or might not affect the information you are after.
The question was how long fixatives can be stored in an TEM lab.
After exhaustive investigation some years ago, we arrived at the following answer: The highest grade glutaraldehyde or paraformaldehyde (distilled and stored in glass vials under inert gas) will deterioate to approximately one half their strengths in 3 weeks assuming they are in a buffer, approximately at pH 7.4, and under continuous refrigeration. Therefore we never keep fixatives for more than one week. We make them up the day of, or the day before we use them. Consider how valuable your sample is and how perfect it is expected to look. A sample which has undergone autolysis will not benefit by ultra-fresh fixation fluids, but living cells in culture will. Oxygen and heat are deleterious to aldehydes. Glutaraldehyde in a mostly empty bottle will not last very long, while glut in a glass vial sealed under inert gas will last many years. Hope this helps. Bye, Hildy
In a similar vein, can any body point me to a vender of such lenses in general. We have a Pixera that we use on the photo tube of several Olympus scopes. We borrowed the screw-in lens from the front of our Dage RS-170 camera. We get the right focal length, but the lens magnification is apparently matched to the size of an RS-170 and not a small CCD chip; therefore we get about an extra three-fold mag over what we see in the eyepieces.
We would like to find a similar lens with a mag of 1x or slightly less. Our adapter lens screws into the front of our Dage or Pixera (whatever you call that kind of mount) and has a 49 mm OD tube that slides into an adapter for our Olympus microscopes.
TIA, Warren
At 02:09 PM 4/3/97 +0000, Nancy wrote: } } Has anyone had experience using a Pixera camera on a TEM? I am } attempting to put one on a Zeiss 902 with a C mount connection. If the camera is } directly mounted, the focal length is not correct. Does anyone know } of an adapter than can be used so theimage can be focused correctly? } } Thanks for any suggestions.
--Boundary (ID jVqXAP1TDFh0bi8LAyJgag) Content-type: TEXT/PLAIN
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Dear Susan,
} } We have recently purchased a Polaron CPD and wanted to hear from } anyone else who has one. I had used one at a University, at which time } once I loaded the specimen chamber and shut the back, opened the fill } valve, the CO2 leaked out the front window. I was concerned and drained } out the CO2. I went to find the instructor, and asked why this was } happening. I assumed a seal was faulty, but she told me that the boat } wasn't loaded properly. The back had closed nicely, and I really didn't } think that it was not loaded properly. Once the chamber was reloaded, } the CO2 tank was empty and I could not finish the run. Once the CO2 } tank was replaced weeks later, I received a phone call saying that the seal } was damaged and that I would have to wait before the new ones came in. } To make a long story short, we now have a Polaron CPD and I am having } the same problem. Sometimes when I load it, it leaks out the front. } Everything seems to be fitting properly, but on occasion, upon filling, it } leaks out the front window. The only reason that I am posting this } question to the listserver and not to the manufacturer, is because another } user thinks that this is normal?!
This is EXACTLY THE SORT OF MATTER TO PUT ON THE LISTSERVER.
Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already.
We have a Polaron CPD 3000 and the seals often leak, particularly the one on the window. I have taken the CPD apart myself to check it. The window seal leaks even though there are no physical defects on it. And you can replace the same seal in the window and next time it will not leak, indicating no permanent damage. So it is not damaged in the sense that a badly loaded boat has pushed up against it and nicked it so it leaks. In fact I can't see how loading the boat would ever make a difference to the integrity of either of the seals, considering how they are located completely inside grooves.
BUT the door seal can be physically damaged by hard particles (say bits of cover slip) getting washed out of the chamber and locating at the sealing surface so they nick the seal as you screw up the door. SO you need to go carefully around the DOOR sealing surface and screw thread with say ethanol on a cotton bud once a week.
AND the seals on the drain valve on the bottom need regular cleaning for the same reason. Grit washes down the drain and scores the sealing surface and O rings in the valve. In fact if you have never done it, go and clean the door seal and drain valve right away. You'll be surprised at the gunge that will be there.
My explanation is to do with seal elasticity. When the CPD is cooled before being pressurised, the neoprene seals lose much of their elasticity and do not expand properly to seal when the chamber is pressurised. So the solvent leaks out and as the seal stays cold it will never seal properly as of course you keep the chamber cold to ensure fast fluid transfer.
The remedy according to Me is to pressurise the CPD BEFORE COOLING IT. BTW we heat and cool the CPD by circulating water through the jacket using a waterbath heater/mixer (Lauda type T, -20deg to 100 deg C.) which has a small centrifugal pump on it and which sits in a 2 Litre stainless tank. At the start we fill up the small tank with ice and add enough water to cover the heater/pump on the mixer. Then we start the pump with the heater off and chill the CPD that way. When we want to warm the CPD we toss away the ice water, replace it with tap water -at- 20 deg or so and turn on the heater which warms the waterbath towards 50 deg.
Notwithstanding all of the above, since the CPD only works if all the seals are good, you must keep a couple of seal kits in your lab ready for quick repairs. Failure of the seals is usually discovered after some poor soul has spent weeks on their experiment and days on processing their tissue only to find the CPD leaks. If you have the capacity to quickly replace any of the seals so their experiment survives your reputation will be much enhanced!
Mel Dickson E.M. Unit, University of NSW Sydney, Australia
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} } Can anyone please comment on the storage properties of common EM/LM fixatives. } We have some older 1% glut/4% formaldehyde in PO4 buffer. Is there some rule } of thumb that people are using, should we analyze before use, are there some } references we can access? } Thank you in advance for your comments. } } Regards, } Ken Baker } } The aldehydes oxidise to acids - formic or glutaric. The reaction is less at lower pH so is accelerated when you buffer the solution to pH7. Our rule is to buffer the fixative just before we use it and discard any buffered fixative older than a week.
Mel Dickson }
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} We solicit your help in: } (1) Sharing knowledge about similar situations
I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off.
The problem will vary according to the soil type between your lab and the new road. Ideally the road would be on swampy ground and your lab on a platfrom carved out of granite. That would give good decoupling. But basically, any effective solution in you laboratory will cost several thousands per instrument, as each instrument will need to be relocated on some heavy support (thick steel plate, thicker concrete slab, mass is what you need) with some very flexible mounts under it (air-springs are ideal). It may be more cost effective to route the road further away.
} (2) Pointing us to the best published sources about EM lab design, } particularly in regard to vibration and preferred distance from nearby roads
} (3) Pointing us to any expert EM lab design firms from whom we } might get information
The classic reference work is "Design of the Electron Microscope Laboratory" by Ronald H Alderson 1975. It is Volume 4 in "Practical Methods in Electron Microscopy" Editor Audrey M Glauert. American Elsevier ISBN 0444 1087 6. Was still available two years back whem I bought my second copy to share with the architect for my new laboratory. Pages 68-86 deal with mechanical vibration and decoupling/damping systems } } Mel Dickson } } } }
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Use a high intensity overhead projector with a cardboard mask to prevent blinding white light.
John D. Warren Area Sales Manager Digital Products Polaroid Corporation
4525 Leonard Parkway Richmond, Virginia 23221-1809
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To any who can help...... I need to find a way to project some 4x5 negatives onto a screen for quantitation purposes. I know that such projectors used to be around, but here at Mayo they are history. I need the projector because I have many low mag micrographs that I am quantitating, and making larger (16x20) prints is not really feasable. Any leads as to where I can borrow, buy or make such a projector would be very helpful!!!
TIA Eugene Krueger GI Research Mayo Foundation krueger.eugene-at-mayo.edu
I've seen this type of "squarish" pattern before. You have most likely lost the central "tip" of your LaB6 filament. It may have fractured off leaving a truncated pyramid shape, which can give this "filament" image. I doubt if there is anything you can do at this point, as you cannot reform the tip if it is missing, however if you are still getting plenty of emission from the LaB6 then you will be okay for normal microscopy. The coherence will likely drop and high resolution imaging may degrade.
Running it "hot" will not reform the tip as you can sometimes do with a Field Emission Source
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Thanks to all those who responded in tracing Edington's Practical electron microscopy book. Tech Books distributes it through their distributor "Ceramic Book and literature Service (CBLS)."
CBLS c an be contacted at 703-758-2539 (ph#)
703-758-1518 (fax#)
or
614-374-9458 (ph#) 614-374-8029 (fax#).
Mohan Kalyanaraman
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Seminar announcement: Optimizing Light Microscopy
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For further details, contact : Dr. Kenneth Piel or Barbara Foster Microscopy/Microscopy Education (MME) 53 Eton Street Springfield, MA 01108 Phone: (413)746-6931 Fax: (413)746-9311 email: mme-at-map.com
To register: Fax or mail the form below to the MME office Name: ___________________________________________________________________ School/Hospital/Company:_________________________________________________Address: ________________________________________________________________ City/State/Zip: _________________________________________________________ Phone: ____________________Fax: _______________email:____________________ Check enclosed (payable to MME) for: _________________ [ ] tuition only [ ] tuition plus lunch [ ] VISA [ ] Master Card Name on credit card: __________________________________________________ Card #: ____________________________________ Expiration date:___________
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... snips } user thinks that this is normal?! Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already. Perhaps it is just not } seated properly. } } P.S. If anyone has recently purchased a Polaron CPD and finds out that } the seal inside the chamber door keeps falling out, a piece of teflon tape } around the seal works wonders! } } } Susan } } } Susan Carbyn } Atlantic Food and Horticulture Research Centre } Agriculture and Agri-Food Canada } Kentville, Nova Scotia B4N 1J5 } Canada } } Phone: (902) 679-5566 } Fax: (902) 679-2311 } E-mail: carbyns-at-em.agr.ca
Well, I wouldn't consider leaking from the from window normal - get the supplier/manufacturer to sort it out.
However, it might be a handling problem rather than an equipment fault. Make sure you go through all the steps slowly, as sudden temperature changes in particular could be causing differential expansion.
I'd also be unhappy about the tape on the door seal. Although unlikely to cause problems, there is a remote chance it might. A better solution is to hold the metal seal edge on and give it a tap against a hard surface - the idea is to make it slightly oval, so it grips its seating.
Regards, Larry Stoter
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Hello Mel and others intetested
Many years ago JEOL News published an article on the design of the EM rooms at the John Innes Institute in the UK. As far as I can recall this dealt in some detail with vibration transmission. We based the design of our EM rooms on this and we have had no vibration problems.
Robin H Cross Director : EM Unit, Rhodes University, Grahamstown, South Africa eurc-at-giraffe.ru.ac.za - tel: +27 461 318168 - fax: +27 461 24377
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Hello Susan
Our Polaron E3000 CPD was bought in 1976 and is still going strong, wuite a lot of use too!
When you say Teflon tape around the seal, I in=magine you mean peripherally rather than through the hole in the middle?!
My experience is that the Doughty/Dowty? seal, the main metal ring plus nitrile (?), tends to split regularly with acetone, every 2-4 months or so. If everything has worked once, then assembly should be ok, it is possible for things to loosen but in my experience that is very unusual. You need to look at the inner face of the seal for any imperfection. I don't know if you get any specail tool, but I made up a steel oblong gizmo which is like a big screwdriver blade whichand gets turned gently with an adjustable wrench.
Hey! I've just realiased, we should have another party for its 21st birthday!!! We get a lot of parties around here, folks!
Best wishes - Keith Ryan Plymouth Marine Lab. UK
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Forwarded message:
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Looking for experienced TEM Service Engineer to service Philips 420 in Kansas City Area. Contact Pete Dondl at sylviapns-at-worldnet.att.net P & S Products, Inc.
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Hi Candace. There was a discussion a while back on this topic which has been archived at the "Tips & Tricks" site. Go to the web address listed at the end of this message and follow the "Tips & Tricks link. Proceed to the Miscl. link and there it is at the top of the list. Good luck.
At 10:07 AM 4/3/97 +0131, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { GO GATORS Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 352-392-1295 ICBR EM Core Lab fax 352-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/ The home of " Tips & Tricks "
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-- [ From: Garber, Charles A. * EMC.Ver #3.1 ] --
In reply to: ================================================== I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off. ================================================== There have been instances where the problem with the rail line has been less due to ground vibrations than to the creation of a transient magnetic field problem that correlated with the passage of a train. In about 1969, there was an SEM installed at a Philadelphia university near the main passenger line of AMRAK and Conrail, and the real problem was more related to the magnetic field (trains were pantograph (electrically) powered) than to vibrations. The problem was ultimately "solved" only by moving the microscope. So don't forget the magnetic field problem potential as well.
Chuck
=================================================== Charles A. Garber, Ph. D. Ph: 1-(610)-436-5400 President 1-(800)-2424-SPI SPI SUPPLIES FAX: 1-(610)-436-5755 PO BOX 656 e-mail: cgarber-at-2spi.com West Chester, PA 19381-0656 USA Cust. Service: spi2spi-at-2spi.com
Look for us! ############################ WWW: http://www.2spi.com ############################ ==================================================
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Dear Susan,
} } We have recently purchased a Polaron CPD and wanted to hear from } anyone else who has one. I had used one at a University, at which time } once I loaded the specimen chamber and shut the back, opened the fill } valve, the CO2 leaked out the front window. I was concerned and drained } out the CO2. I went to find the instructor, and asked why this was } happening. I assumed a seal was faulty, but she told me that the boat } wasn't loaded properly. The back had closed nicely, and I really didn't } think that it was not loaded properly. Once the chamber was reloaded, } the CO2 tank was empty and I could not finish the run. Once the CO2 } tank was replaced weeks later, I received a phone call saying that the seal } was damaged and that I would have to wait before the new ones came in. } To make a long story short, we now have a Polaron CPD and I am having } the same problem. Sometimes when I load it, it leaks out the front. } Everything seems to be fitting properly, but on occasion, upon filling, it } leaks out the front window. The only reason that I am posting this } question to the listserver and not to the manufacturer, is because another } user thinks that this is normal?!
This is EXACTLY THE SORT OF MATTER TO PUT ON THE LISTSERVER.
Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already.
We have a Polaron CPD 3000 and the seals often leak, particularly the one on the window. I have taken the CPD apart myself to check it. The window seal leaks even though there are no physical defects on it. And you can replace the same seal in the window and next time it will not leak, indicating no permanent damage. So it is not damaged in the sense that a badly loaded boat has pushed up against it and nicked it so it leaks. In fact I can't see how loading the boat would ever make a difference to the integrity of either of the seals, considering how they are located completely inside grooves.
BUT the door seal can be physically damaged by hard particles (say bits of cover slip) getting washed out of the chamber and locating at the sealing surface so they nick the seal as you screw up the door. SO you need to go carefully around the DOOR sealing surface and screw thread with say ethanol on a cotton bud once a week.
AND the seals on the drain valve on the bottom need regular cleaning for the same reason. Grit washes down the drain and scores the sealing surface and O rings in the valve. In fact if you have never done it, go and clean the door seal and drain valve right away. You'll be surprised at the gunge that will be there.
My explanation is to do with seal elasticity. When the CPD is cooled before being pressurised, the neoprene seals lose much of their elasticity and do not expand properly to seal when the chamber is pressurised. So the solvent leaks out and as the seal stays cold it will never seal properly as of course you keep the chamber cold to ensure fast fluid transfer.
The remedy according to Me is to pressurise the CPD BEFORE COOLING IT. BTW we heat and cool the CPD by circulating water through the jacket using a waterbath heater/mixer (Lauda type T, -20deg to 100 deg C.) which has a small centrifugal pump on it and which sits in a 2 Litre stainless tank. At the start we fill up the small tank with ice and add enough water to cover the heater/pump on the mixer. Then we start the pump with the heater off and chill the CPD that way. When we want to warm the CPD we toss away the ice water, replace it with tap water -at- 20 deg or so and turn on the heater which warms the waterbath towards 50 deg.
Notwithstanding all of the above, since the CPD only works if all the seals are good, you must keep a couple of seal kits in your lab ready for quick repairs. Failure of the seals is usually discovered after some poor soul has spent weeks on their experiment and days on processing their tissue only to find the CPD leaks. If you have the capacity to quickly replace any of the seals so their experiment survives your reputation will be much enhanced!
Mel Dickson E.M. Unit, University of NSW Sydney, Australia
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} } Can anyone please comment on the storage properties of common EM/LM fixatives. } We have some older 1% glut/4% formaldehyde in PO4 buffer. Is there some rule } of thumb that people are using, should we analyze before use, are there some } references we can access? } Thank you in advance for your comments. } } Regards, } Ken Baker } } The aldehydes oxidise to acids - formic or glutaric. The reaction is less at lower pH so is accelerated when you buffer the solution to pH7. Our rule is to buffer the fixative just before we use it and discard any buffered fixative older than a week.
Mel Dickson }
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} We solicit your help in: } (1) Sharing knowledge about similar situations
I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off.
The problem will vary according to the soil type between your lab and the new road. Ideally the road would be on swampy ground and your lab on a platfrom carved out of granite. That would give good decoupling. But basically, any effective solution in you laboratory will cost several thousands per instrument, as each instrument will need to be relocated on some heavy support (thick steel plate, thicker concrete slab, mass is what you need) with some very flexible mounts under it (air-springs are ideal). It may be more cost effective to route the road further away.
} (2) Pointing us to the best published sources about EM lab design, } particularly in regard to vibration and preferred distance from nearby roads
} (3) Pointing us to any expert EM lab design firms from whom we } might get information
The classic reference work is "Design of the Electron Microscope Laboratory" by Ronald H Alderson 1975. It is Volume 4 in "Practical Methods in Electron Microscopy" Editor Audrey M Glauert. American Elsevier ISBN 0444 1087 6. Was still available two years back whem I bought my second copy to share with the architect for my new laboratory. Pages 68-86 deal with mechanical vibration and decoupling/damping systems } } Mel Dickson } } } }
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Dear Angus,
Potassium hydroxide works well for etching silicon without attacking silicon dioxide. It might also work well with the nitride as an etch stop, but I don't have any experience with that. Concentration and temperature could be varied to enhance the selectivity for Si vs nitride.
D. Clark Turner MOXTEK, Inc. 452 West 1260 North Orem, Utah 84057 (801) 225-0930 email moxtek-at-moxtek.win.net
} Dear All,
} Does anyone know of a selective etch that will etch away silicon } but not silicon nitride? I'm looking at a multilayer sample in plan } view for TEM and hope to back etch the si substrate and use the } nitride as a stop layer.
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-- [ From: Garber, Charles A. * EMC.Ver #3.1 ] --
In reply to: ================================================== I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off. ================================================== There have been instances where the problem with the rail line has been less due to ground vibrations than to the creation of a transient magnetic field problem that correlated with the passage of a train. In about 1969, there was an SEM installed at a Philadelphia university near the main passenger line of AMRAK and Conrail, and the real problem was more related to the magnetic field (trains were pantograph (electrically) powered) than to vibrations. The problem was ultimately "solved" only by moving the microscope. So don't forget the magnetic field problem potential as well.
Chuck
=================================================== Charles A. Garber, Ph. D. Ph: 1-(610)-436-5400 President 1-(800)-2424-SPI SPI SUPPLIES FAX: 1-(610)-436-5755 PO BOX 656 e-mail: cgarber-at-2spi.com West Chester, PA 19381-0656 USA Cust. Service: spi2spi-at-2spi.com
Look for us! ############################ WWW: http://www.2spi.com ############################ ==================================================
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Hi Candace. There was a discussion a while back on this topic which has been archived at the "Tips & Tricks" site. Go to the web address listed at the end of this message and follow the "Tips & Tricks link. Proceed to the Miscl. link and there it is at the top of the list. Good luck.
At 10:07 AM 4/3/97 +0131, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { GO GATORS Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 352-392-1295 ICBR EM Core Lab fax 352-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/ The home of " Tips & Tricks "
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Assuming that not the entire film area is required, the alternative is to make contact prints of the required areas. 35mm ortho film or if you rather use a slightly larger format, cut up TEM sheetfilm to suit super 35 mm mounts. Make sure the copying is done emulsion to emulsion and use an enlarger as your lightsource. Once the method is established its quite fast. Jim Darley
ProSciTech Microscopy PLUS PO Box 111, Thuringowa QLD 4817 Australia Phone +61 77 740 370 Fax: +61 77 892 313 Great microscopy catalogue, 300+ Links, MSDS ************************ http://www.proscitech.com.au
} I need to find a way to project some 4x5 negatives onto a screen for } quantitation purposes. I need the projector because I have many low } mag micrographs that I am quantitating, and making larger (16x20) prints is } not really feasable. } Eugene Krueger } GI Research } Mayo Foundation } krueger.eugene-at-mayo.edu
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Looking for experienced TEM Service Engineer to service Philips 420 in Kansas City Area. Contact Pete Dondl at sylviapns-at-worldnet.att.net P & S Products, Inc.
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Dear Susan,
} } We have recently purchased a Polaron CPD and wanted to hear from } anyone else who has one. I had used one at a University, at which time } once I loaded the specimen chamber and shut the back, opened the fill } valve, the CO2 leaked out the front window. I was concerned and drained } out the CO2. I went to find the instructor, and asked why this was } happening. I assumed a seal was faulty, but she told me that the boat } wasn't loaded properly. The back had closed nicely, and I really didn't } think that it was not loaded properly. Once the chamber was reloaded, } the CO2 tank was empty and I could not finish the run. Once the CO2 } tank was replaced weeks later, I received a phone call saying that the seal } was damaged and that I would have to wait before the new ones came in. } To make a long story short, we now have a Polaron CPD and I am having } the same problem. Sometimes when I load it, it leaks out the front. } Everything seems to be fitting properly, but on occasion, upon filling, it } leaks out the front window. The only reason that I am posting this } question to the listserver and not to the manufacturer, is because another } user thinks that this is normal?!
This is EXACTLY THE SORT OF MATTER TO PUT ON THE LISTSERVER.
Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already.
We have a Polaron CPD 3000 and the seals often leak, particularly the one on the window. I have taken the CPD apart myself to check it. The window seal leaks even though there are no physical defects on it. And you can replace the same seal in the window and next time it will not leak, indicating no permanent damage. So it is not damaged in the sense that a badly loaded boat has pushed up against it and nicked it so it leaks. In fact I can't see how loading the boat would ever make a difference to the integrity of either of the seals, considering how they are located completely inside grooves.
BUT the door seal can be physically damaged by hard particles (say bits of cover slip) getting washed out of the chamber and locating at the sealing surface so they nick the seal as you screw up the door. SO you need to go carefully around the DOOR sealing surface and screw thread with say ethanol on a cotton bud once a week.
AND the seals on the drain valve on the bottom need regular cleaning for the same reason. Grit washes down the drain and scores the sealing surface and O rings in the valve. In fact if you have never done it, go and clean the door seal and drain valve right away. You'll be surprised at the gunge that will be there.
My explanation is to do with seal elasticity. When the CPD is cooled before being pressurised, the neoprene seals lose much of their elasticity and do not expand properly to seal when the chamber is pressurised. So the solvent leaks out and as the seal stays cold it will never seal properly as of course you keep the chamber cold to ensure fast fluid transfer.
The remedy according to Me is to pressurise the CPD BEFORE COOLING IT. BTW we heat and cool the CPD by circulating water through the jacket using a waterbath heater/mixer (Lauda type T, -20deg to 100 deg C.) which has a small centrifugal pump on it and which sits in a 2 Litre stainless tank. At the start we fill up the small tank with ice and add enough water to cover the heater/pump on the mixer. Then we start the pump with the heater off and chill the CPD that way. When we want to warm the CPD we toss away the ice water, replace it with tap water -at- 20 deg or so and turn on the heater which warms the waterbath towards 50 deg.
Notwithstanding all of the above, since the CPD only works if all the seals are good, you must keep a couple of seal kits in your lab ready for quick repairs. Failure of the seals is usually discovered after some poor soul has spent weeks on their experiment and days on processing their tissue only to find the CPD leaks. If you have the capacity to quickly replace any of the seals so their experiment survives your reputation will be much enhanced!
Mel Dickson E.M. Unit, University of NSW Sydney, Australia
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} } Can anyone please comment on the storage properties of common EM/LM fixatives. } We have some older 1% glut/4% formaldehyde in PO4 buffer. Is there some rule } of thumb that people are using, should we analyze before use, are there some } references we can access? } Thank you in advance for your comments. } } Regards, } Ken Baker } } The aldehydes oxidise to acids - formic or glutaric. The reaction is less at lower pH so is accelerated when you buffer the solution to pH7. Our rule is to buffer the fixative just before we use it and discard any buffered fixative older than a week.
Mel Dickson }
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} We solicit your help in: } (1) Sharing knowledge about similar situations
I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off.
The problem will vary according to the soil type between your lab and the new road. Ideally the road would be on swampy ground and your lab on a platfrom carved out of granite. That would give good decoupling. But basically, any effective solution in you laboratory will cost several thousands per instrument, as each instrument will need to be relocated on some heavy support (thick steel plate, thicker concrete slab, mass is what you need) with some very flexible mounts under it (air-springs are ideal). It may be more cost effective to route the road further away.
} (2) Pointing us to the best published sources about EM lab design, } particularly in regard to vibration and preferred distance from nearby roads
} (3) Pointing us to any expert EM lab design firms from whom we } might get information
The classic reference work is "Design of the Electron Microscope Laboratory" by Ronald H Alderson 1975. It is Volume 4 in "Practical Methods in Electron Microscopy" Editor Audrey M Glauert. American Elsevier ISBN 0444 1087 6. Was still available two years back whem I bought my second copy to share with the architect for my new laboratory. Pages 68-86 deal with mechanical vibration and decoupling/damping systems } } Mel Dickson } } } }
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Thanks to all those who responded in tracing Edington's Practical electron microscopy book. Tech Books distributes it through their distributor "Ceramic Book and literature Service (CBLS)."
CBLS c an be contacted at 703-758-2539 (ph#)
703-758-1518 (fax#)
or
614-374-9458 (ph#) 614-374-8029 (fax#).
Mohan Kalyanaraman
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Dear Susan,
} } We have recently purchased a Polaron CPD and wanted to hear from } anyone else who has one. I had used one at a University, at which time } once I loaded the specimen chamber and shut the back, opened the fill } valve, the CO2 leaked out the front window. I was concerned and drained } out the CO2. I went to find the instructor, and asked why this was } happening. I assumed a seal was faulty, but she told me that the boat } wasn't loaded properly. The back had closed nicely, and I really didn't } think that it was not loaded properly. Once the chamber was reloaded, } the CO2 tank was empty and I could not finish the run. Once the CO2 } tank was replaced weeks later, I received a phone call saying that the seal } was damaged and that I would have to wait before the new ones came in. } To make a long story short, we now have a Polaron CPD and I am having } the same problem. Sometimes when I load it, it leaks out the front. } Everything seems to be fitting properly, but on occasion, upon filling, it } leaks out the front window. The only reason that I am posting this } question to the listserver and not to the manufacturer, is because another } user thinks that this is normal?!
This is EXACTLY THE SORT OF MATTER TO PUT ON THE LISTSERVER.
Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already.
We have a Polaron CPD 3000 and the seals often leak, particularly the one on the window. I have taken the CPD apart myself to check it. The window seal leaks even though there are no physical defects on it. And you can replace the same seal in the window and next time it will not leak, indicating no permanent damage. So it is not damaged in the sense that a badly loaded boat has pushed up against it and nicked it so it leaks. In fact I can't see how loading the boat would ever make a difference to the integrity of either of the seals, considering how they are located completely inside grooves.
BUT the door seal can be physically damaged by hard particles (say bits of cover slip) getting washed out of the chamber and locating at the sealing surface so they nick the seal as you screw up the door. SO you need to go carefully around the DOOR sealing surface and screw thread with say ethanol on a cotton bud once a week.
AND the seals on the drain valve on the bottom need regular cleaning for the same reason. Grit washes down the drain and scores the sealing surface and O rings in the valve. In fact if you have never done it, go and clean the door seal and drain valve right away. You'll be surprised at the gunge that will be there.
My explanation is to do with seal elasticity. When the CPD is cooled before being pressurised, the neoprene seals lose much of their elasticity and do not expand properly to seal when the chamber is pressurised. So the solvent leaks out and as the seal stays cold it will never seal properly as of course you keep the chamber cold to ensure fast fluid transfer.
The remedy according to Me is to pressurise the CPD BEFORE COOLING IT. BTW we heat and cool the CPD by circulating water through the jacket using a waterbath heater/mixer (Lauda type T, -20deg to 100 deg C.) which has a small centrifugal pump on it and which sits in a 2 Litre stainless tank. At the start we fill up the small tank with ice and add enough water to cover the heater/pump on the mixer. Then we start the pump with the heater off and chill the CPD that way. When we want to warm the CPD we toss away the ice water, replace it with tap water -at- 20 deg or so and turn on the heater which warms the waterbath towards 50 deg.
Notwithstanding all of the above, since the CPD only works if all the seals are good, you must keep a couple of seal kits in your lab ready for quick repairs. Failure of the seals is usually discovered after some poor soul has spent weeks on their experiment and days on processing their tissue only to find the CPD leaks. If you have the capacity to quickly replace any of the seals so their experiment survives your reputation will be much enhanced!
Mel Dickson E.M. Unit, University of NSW Sydney, Australia
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} } Can anyone please comment on the storage properties of common EM/LM fixatives. } We have some older 1% glut/4% formaldehyde in PO4 buffer. Is there some rule } of thumb that people are using, should we analyze before use, are there some } references we can access? } Thank you in advance for your comments. } } Regards, } Ken Baker } } The aldehydes oxidise to acids - formic or glutaric. The reaction is less at lower pH so is accelerated when you buffer the solution to pH7. Our rule is to buffer the fixative just before we use it and discard any buffered fixative older than a week.
Mel Dickson }
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} We solicit your help in: } (1) Sharing knowledge about similar situations
I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off.
The problem will vary according to the soil type between your lab and the new road. Ideally the road would be on swampy ground and your lab on a platfrom carved out of granite. That would give good decoupling. But basically, any effective solution in you laboratory will cost several thousands per instrument, as each instrument will need to be relocated on some heavy support (thick steel plate, thicker concrete slab, mass is what you need) with some very flexible mounts under it (air-springs are ideal). It may be more cost effective to route the road further away.
} (2) Pointing us to the best published sources about EM lab design, } particularly in regard to vibration and preferred distance from nearby roads
} (3) Pointing us to any expert EM lab design firms from whom we } might get information
The classic reference work is "Design of the Electron Microscope Laboratory" by Ronald H Alderson 1975. It is Volume 4 in "Practical Methods in Electron Microscopy" Editor Audrey M Glauert. American Elsevier ISBN 0444 1087 6. Was still available two years back whem I bought my second copy to share with the architect for my new laboratory. Pages 68-86 deal with mechanical vibration and decoupling/damping systems } } Mel Dickson } } } }
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A Lab6 cathode running on a Hitachi H7100 TEM (manufacturers brand) developed a bright rectanglar-shaped emmision pattern over the last 50-100 hours of operation, although it can be biased back to a small circular spot. We hadnt seen this before. The tip of the cathode shows a distinctly bar-shaped tip when viewed in a light microscope.
The cathode has been very stable over its 450 hours of life, although it seems to move a little vertically as it heats up. There is plenty of Lab6 left.
Does anyone know what causes the pattern, and what are the chances of getting back to a squarish/maltese cross type image if we, say, run it a bit hot for a while?
regards, Sally ---------------------------------------------------------------------- Sally Stowe |Email: stowe-at-rsbs.anu.edu.au Facility Coordinator |Post: ANU Electron Microscopy Unit |ANUEMU (RSBS) Ph 61 6 249 2743 |Australian National Univ. FAX 61 6 249 4891 |Canberra, http://online.anu.edu.au/EMU/home.htm |AUSTRALIA 0200
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A Lab6 cathode running on a Hitachi H7100 TEM (manufacturers brand) developed a bright rectanglar-shaped emmision pattern over the last 50-100 hours of operation, although it can be biased back to a small circular spot. We hadnt seen this before. The tip of the cathode shows a distinctly bar-shaped tip when viewed in a light microscope.
The cathode has been very stable over its 450 hours of life, although it seems to move a little vertically as it heats up. There is plenty of Lab6 left.
Does anyone know what causes the pattern, and what are the chances of getting back to a squarish/maltese cross type image if we, say, run it a bit hot for a while?
regards, Sally ---------------------------------------------------------------------- Sally Stowe |Email: stowe-at-rsbs.anu.edu.au Facility Coordinator |Post: ANU Electron Microscopy Unit |ANUEMU (RSBS) Ph 61 6 249 2743 |Australian National Univ. FAX 61 6 249 4891 |Canberra, http://online.anu.edu.au/EMU/home.htm |AUSTRALIA 0200
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... snips } user thinks that this is normal?! Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already. Perhaps it is just not } seated properly. } } P.S. If anyone has recently purchased a Polaron CPD and finds out that } the seal inside the chamber door keeps falling out, a piece of teflon tape } around the seal works wonders! } } } Susan } } } Susan Carbyn } Atlantic Food and Horticulture Research Centre } Agriculture and Agri-Food Canada } Kentville, Nova Scotia B4N 1J5 } Canada } } Phone: (902) 679-5566 } Fax: (902) 679-2311 } E-mail: carbyns-at-em.agr.ca
Well, I wouldn't consider leaking from the from window normal - get the supplier/manufacturer to sort it out.
However, it might be a handling problem rather than an equipment fault. Make sure you go through all the steps slowly, as sudden temperature changes in particular could be causing differential expansion.
I'd also be unhappy about the tape on the door seal. Although unlikely to cause problems, there is a remote chance it might. A better solution is to hold the metal seal edge on and give it a tap against a hard surface - the idea is to make it slightly oval, so it grips its seating.
Regards, Larry Stoter
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Hello Mel and others intetested
Many years ago JEOL News published an article on the design of the EM rooms at the John Innes Institute in the UK. As far as I can recall this dealt in some detail with vibration transmission. We based the design of our EM rooms on this and we have had no vibration problems.
Robin H Cross Director : EM Unit, Rhodes University, Grahamstown, South Africa eurc-at-giraffe.ru.ac.za - tel: +27 461 318168 - fax: +27 461 24377
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A Lab6 cathode running on a Hitachi H7100 TEM (manufacturers brand) developed a bright rectanglar-shaped emmision pattern over the last 50-100 hours of operation, although it can be biased back to a small circular spot. We hadnt seen this before. The tip of the cathode shows a distinctly bar-shaped tip when viewed in a light microscope.
The cathode has been very stable over its 450 hours of life, although it seems to move a little vertically as it heats up. There is plenty of Lab6 left.
Does anyone know what causes the pattern, and what are the chances of getting back to a squarish/maltese cross type image if we, say, run it a bit hot for a while?
regards, Sally ---------------------------------------------------------------------- Sally Stowe |Email: stowe-at-rsbs.anu.edu.au Facility Coordinator |Post: ANU Electron Microscopy Unit |ANUEMU (RSBS) Ph 61 6 249 2743 |Australian National Univ. FAX 61 6 249 4891 |Canberra, http://online.anu.edu.au/EMU/home.htm |AUSTRALIA 0200
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Hello Susan
Our Polaron E3000 CPD was bought in 1976 and is still going strong, wuite a lot of use too!
When you say Teflon tape around the seal, I in=magine you mean peripherally rather than through the hole in the middle?!
My experience is that the Doughty/Dowty? seal, the main metal ring plus nitrile (?), tends to split regularly with acetone, every 2-4 months or so. If everything has worked once, then assembly should be ok, it is possible for things to loosen but in my experience that is very unusual. You need to look at the inner face of the seal for any imperfection. I don't know if you get any specail tool, but I made up a steel oblong gizmo which is like a big screwdriver blade whichand gets turned gently with an adjustable wrench.
Hey! I've just realiased, we should have another party for its 21st birthday!!! We get a lot of parties around here, folks!
Best wishes - Keith Ryan Plymouth Marine Lab. UK
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Forwarded message:
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Thanks to all those who responded in tracing Edington's Practical electron microscopy book. Tech Books distributes it through their distributor "Ceramic Book and literature Service (CBLS)."
CBLS c an be contacted at 703-758-2539 (ph#)
703-758-1518 (fax#)
or
614-374-9458 (ph#) 614-374-8029 (fax#).
Mohan Kalyanaraman
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-- [ From: Blackwood, Andy * EMC.Ver #3.0 ] --
3 April 1997
Greetings, John Bozzola:
I saw your posting about the JEOL sample a few days ago, and since nobody else seems to have made the point, I thought that a few comments might help. My bias is rather obvious--as Laboratory Director of Structure Probe, Inc., I am responsible for the production of the SPI Supplies reference samples.
We offer three different samples, because our customers request various sizes. All are the same gold on carbon construction, and they offer sharp contrast between gold (high emission of secondary electrons) and carbon (low emission of secondary electrons). We characterize the samples by the average size of the particles (small is around 10 nm, medium is around 30 nm and large is around 100 nm). The actual reference, however, is the gap between gold particles, and by looking around a little, you can find any size gap you wish to use on any of the samples. Larger laboratories have obtained a set of three samples to cover the entire range. Details may be found on our web site.
It certainly is possible to set up to produce such samples in your own laboratory, but by the time you've finished fine tuning the "art" for your local conditions, and then verified that what you made is what you hoped to make, you may conclude that it would have been easier to obtain a commercially made sample.
Andy
Andrew W. Blackwood, Ph.D. Structure Probe, Inc. P.O. Box 656 West Chester, PA 19381-0656 Ph: 1 610 436 5400 FAX: 1 610 436 5755 e-mail: ablackwood-at-2spi.com WWW: http://www.2spi.com
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Looking for experienced TEM Service Engineer to service Philips 420 in Kansas City Area. Contact Pete Dondl at sylviapns-at-worldnet.att.net P & S Products, Inc.
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-- [ From: Garber, Charles A. * EMC.Ver #3.1 ] --
In reply to: ================================================== I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off. ================================================== There have been instances where the problem with the rail line has been less due to ground vibrations than to the creation of a transient magnetic field problem that correlated with the passage of a train. In about 1969, there was an SEM installed at a Philadelphia university near the main passenger line of AMRAK and Conrail, and the real problem was more related to the magnetic field (trains were pantograph (electrically) powered) than to vibrations. The problem was ultimately "solved" only by moving the microscope. So don't forget the magnetic field problem potential as well.
Chuck
=================================================== Charles A. Garber, Ph. D. Ph: 1-(610)-436-5400 President 1-(800)-2424-SPI SPI SUPPLIES FAX: 1-(610)-436-5755 PO BOX 656 e-mail: cgarber-at-2spi.com West Chester, PA 19381-0656 USA Cust. Service: spi2spi-at-2spi.com
Look for us! ############################ WWW: http://www.2spi.com ############################ ==================================================
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Hi Candace. There was a discussion a while back on this topic which has been archived at the "Tips & Tricks" site. Go to the web address listed at the end of this message and follow the "Tips & Tricks link. Proceed to the Miscl. link and there it is at the top of the list. Good luck.
At 10:07 AM 4/3/97 +0131, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { GO GATORS Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 352-392-1295 ICBR EM Core Lab fax 352-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/ The home of " Tips & Tricks "
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On Fri, 4 Apr 1997 sylviapns-at-worldnet.att.net wrote:
} Date: Fri, 04 Apr 1997 11:12 -0500 (EST) } From: sylviapns-at-worldnet.att.net } To: MICROSCOPY-at-Sparc5.Microscopy.Com } Subject: TEM Service } } } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Looking for experienced TEM Service Engineer to service Philips 420 in } Kansas City Area. } Contact } Pete Dondl at sylviapns-at-worldnet.att.net } P & S Products, Inc. } Call Philips: 1 800 432 1PEI}
Sara E. Miller, Ph. D. P. O. Box 3020 Duke University Medical Center Durham, NC 27710 Ph: 919 684-3452 FAX: 919 684-8735
Another interesting problem I was told about related to a EM installed in Eastern Europe a number of years back. There were continual, intermittent resolution and noise problems, worse during the day, not too often at night and after midnight, the problems disappeared.
After a lot of investigations, the problems was finally traced to the local trams. What was happening was that everything was OK until the trams reached a nearby hill - the extra load in pulling up the hill (trams going down the hill weren't a problem) dragged the mains supply voltage below an acceptable level and caused a whole series of instabilities in the EM.
Regards,
--------------------------------------------------------------- Dr L. P. Stoter Technical Editor, MICROSCOPY & ANALYSIS Technesis 17, Rocks Park Road email: LPS-at-teknesis.demon.co.uk Uckfield, E. Sussex Phone: +44 (0)1825 766911 TN22 2AT Fax: +44 (0)1825 766911 United Kingdom ---------------------------------------------------------------
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Postdoctoral Positions in Biophysics, Biochemistry and Biomathematics.
Two NSF-funded fellowships are available immediately for interdisciplinary studies of macromolecular assemblies in the following areas:
1. Computational analysis of atomic interactions: including development of holistic mathematical analysis of fundamental interactions, exploiting the complementarity of data from diffraction, NMR spectroscopy and electron microscopy. Candidates should have backgrounds in computational methods development in crystallography, NMR, EM or computational structural biology, or training in the mathematical / physical sciences combined with demonstrable interest in structural biology.
2. Structure & assembly of macromolecular complexes of fibroblast growth factor & receptor: the candidate will pursue structural analyses of soluble forms of fibroblast growth factor receptors and receptor/growth factor complexes. Candidates should have a background in x-ray crystallography, protein purification and molecular biology. Experience in baculovirus expression systems, and cell culture would be a plus.
3. Comparing protein motion in solution and the crystal. The fellow will develop an approach to completely map the dynamics of the C-terminal domain of the diptheria toxin repressor protein by combining solution NMR spectroscopy and diffuse-scatter X-ray crystallography. Candidates should have a strong background in NMR spectroscopy or x-ray crystallography, with an interest in protein dynamics.
4. Protein dynamics as measured by EPR and X-ray diffraction methods: the candidate will develop "rules" governing the dynamics of spin labels attached to proteins of known structure. The dynamics observed in the spin labeled crystals using EPR will be compared to the diffuse scatter and the temperature factors from x-ray studies. The candidate should have background in magnetic resonance techniques or x-ray crystallography.
Consistent with these interdisciplinary fellowships, trainees will be mentored jointly by faculty from at least two fields: Michael Blaber, Don Caspar and Michael Chapman (crystallography), Tim Cross and Tim Logan (NMR), Piotr Fajer (EPR) and Ken Taylor (EM). Supplementing experimental facilities at the Institute of Molecular Biophysics, close ties are enjoyed with the National High Field Magnetic Laboratory and the Supercomputer Computation Research Institute on campus. Fellows will enjoy a stimulating intellectual environment, pleasant weather, national forests and pristine gulf beaches.
Candidates should send a resume and arrange for 3 letters of reference to be forwarded to the Institute of Molecular Biophysics, Attn: Carolyn Moore / Post-doc. Fellows, Florida State University, Tallahassee, FL 32306-3015, USA, prior to July 1st. The project of interest should be stated at the top of the resume. Additional information about the Structural Biology Program can be found at http://www.sb.fsu.edu/.
-- Michael S. Chapman (chapman-at-sb.fsu.edu) http://www.sb.fsu.edu/~chapman/ Assistant Prof. Chemistry (904) 644-8354 FAX: (904) 644-3257 Institute of Molecular Biophysics, Florida State University, Tallahassee, FL 32306-3015
Speaking of vibrations, can anyone provide information (name, address, phone, FAX, e-mail, www address, etc.) for vendors of vibration isolation pads and platforms. I have a new stereo microscope installation that is being bothered by building vibrations.
Thanks...
Larry Sutter Michigan Technological University Dept. of Civil and Environmental Engineering 1400 Townsend Dr. Houghton, MI 49931
At 06:30 PM 06-04-97 -0400, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
You can do it yourself.
Get a 2 ft (or so) square of paving cement from a garden supply (or hardware ) company. Buy 4 tennis balls. Put the slab on some sturdy bench with a tennis ball under each corner. Put the microscope on the slab. For a more compliant isolator use a small inner tube from some small wheel.
The problem does not appear to originate at the Microscopy Listserver. It looks to be a computer/email system at DUKE.EDU that is redirecting mail back to the server. For the moment I have removed the following addresses from the subscription list. If this cures the problem we know that the problem has been at least isolated.
One of my colleagues asks: } } Does anyone know where I can get a program that can calculate } the multi-beam diffraction contrast of a (given) strain field? } (i.e. not just a two-beam calculation)
TIA,
Stephen. ............................................................... : Stephen Anderson Australian Key Centre for : : Microscopy and Microanalysis : : Email stephen-at-emu.usyd.edu.au The University of Sydney : : Telephone (+61)-2-9351 7552 NSW 2006 : : Facsimile (+61)-2-9351 7682 Australia : :.............................................................:
We sometimes store small aliquots of fixative at minus 20 C. I'm not sure this is effective but it seems like a reasonable thing to do. Seems as though I came upon this in one of the early editions of Hyats general EM techniques book. ******************************************************* G.W. Erdos, Ph.D. Phone: 352-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax: 352-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 http://www.biotech.ufl.edu/~emcl/ Home of the #1 Gators ***** "Many shall run to and fro, and knowledge shall be increased" Daniel 12:4
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Colleagues...
The Microscopy Node was down for most of the day today. There was another problem with Ameritek. It must be all that warm weather we are having in Chicago ;-)
As for the problem about the duplicate postings as far as I can tell the problem does not appear to originate at the Microscopy Listserver. It looks to be a computer/email system at DUKE.EDU that is redirecting mail back to the server. For the moment I have removed the following addresses from the subscription list. If this cures the problem we know that the problem has been at least isolated.
there is a program called SIMCON that will probably do what you want. It's written at the materials department of the University of Leuven by Koen Janssens. You'll find a reference for it in "Ultramicroscopy 45, p. 323 (1992)" and at "http://www.mtm.kuleuven.ac.be/~simcon/". The author has now left the group and works at OCAS, J.F. Kennedylaan 3, B-9060 Zelzate.
1. When a gold coated specimen is viewed in SEM at ~10k or higher, a fine structure of gold coating becomes visible. Are there optimal conditions of sputter coating (sputter current, time, distance target-specimen, argon pressure, other gas than argon) to minimise the artefact?
2. What is Au-Pd target? I thought that it was just an alloy one, but a supplier of the coater says that using the alloy target is not enough, that a coater with simultaneous sputtering Au and Pd from two different targets is required to produce continuous coating.
Of course, it's better to get a FEG and use low voltage, but I want to do it with a magnetron sputter coater and conventional JSM 6400.
Thank you,
Alex
__________________ Alexander Titkov
Millennium Inorganic Chemicals PO Box 245 Bunbury WA 6231 Australia Ph (097) 808 505 FAX: (097) 808 444 E-mail: scm!atitkov-at-scmaust.attmail.com
This last piece of information was passed along to me by the Manufacturer of the Polaron CPD.
Please ammend the e-mail or notify the customer with the faulty CPD that the seals used were not in fact made by DOWTY but of similar design, from another manufacturer, these in fact were of the wrong material so in practice the material was not faulty but the manufactured item itself was below spec.
Susan
Susan Carbyn Atlantic Food and Horticulture Research Station Agriculture and Agri-Food Canada Kentville, Nova Scotia B4N 1J5 Canada
Meeting: Spring Meeting of the Appalachian REgional Microscopy Society Dates: April 17 & 18, 1997 Topic: Registration at Comfort Suites, Noon to 5pm, Thursday, 4/17/97 Workshops 1pm to 5pm, Thursday, 4/17/97 Social and Banquet, 6pm, Thursday, 4/17/97 Technical Presentations at Enka Lake Club, 8am to 1pm, Friday, 4/18/97 Sponsor: BASF, Enka, NC Location: Asheville, NC, USA Interests: Both Physical & Biological Sciences Fields: Light/Optical, SEM, TEM Contact: Susan Read, BASF Corporation, Sand Hill Road, Enka, NC, 28728, USA Tel: (704)667-6353 Fax: (704)667-6903 E-mail: reads-at-basf.com WWW: http://www.clinck.edu/~jrb/arems.html ---------------------------------------------------------------------------
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Dear Susan,
} } We have recently purchased a Polaron CPD and wanted to hear from } anyone else who has one. I had used one at a University, at which time } once I loaded the specimen chamber and shut the back, opened the fill } valve, the CO2 leaked out the front window. I was concerned and drained } out the CO2. I went to find the instructor, and asked why this was } happening. I assumed a seal was faulty, but she told me that the boat } wasn't loaded properly. The back had closed nicely, and I really didn't } think that it was not loaded properly. Once the chamber was reloaded, } the CO2 tank was empty and I could not finish the run. Once the CO2 } tank was replaced weeks later, I received a phone call saying that the seal } was damaged and that I would have to wait before the new ones came in. } To make a long story short, we now have a Polaron CPD and I am having } the same problem. Sometimes when I load it, it leaks out the front. } Everything seems to be fitting properly, but on occasion, upon filling, it } leaks out the front window. The only reason that I am posting this } question to the listserver and not to the manufacturer, is because another } user thinks that this is normal?!
This is EXACTLY THE SORT OF MATTER TO PUT ON THE LISTSERVER.
Even if the boat was not fitting properly, } should the vessel still leak out the front? I would appreciate any } enlightment on this problem. It is a brandnew CPD and thus I have my } doubts that the seal would be damaged already.
We have a Polaron CPD 3000 and the seals often leak, particularly the one on the window. I have taken the CPD apart myself to check it. The window seal leaks even though there are no physical defects on it. And you can replace the same seal in the window and next time it will not leak, indicating no permanent damage. So it is not damaged in the sense that a badly loaded boat has pushed up against it and nicked it so it leaks. In fact I can't see how loading the boat would ever make a difference to the integrity of either of the seals, considering how they are located completely inside grooves.
BUT the door seal can be physically damaged by hard particles (say bits of cover slip) getting washed out of the chamber and locating at the sealing surface so they nick the seal as you screw up the door. SO you need to go carefully around the DOOR sealing surface and screw thread with say ethanol on a cotton bud once a week.
AND the seals on the drain valve on the bottom need regular cleaning for the same reason. Grit washes down the drain and scores the sealing surface and O rings in the valve. In fact if you have never done it, go and clean the door seal and drain valve right away. You'll be surprised at the gunge that will be there.
My explanation is to do with seal elasticity. When the CPD is cooled before being pressurised, the neoprene seals lose much of their elasticity and do not expand properly to seal when the chamber is pressurised. So the solvent leaks out and as the seal stays cold it will never seal properly as of course you keep the chamber cold to ensure fast fluid transfer.
The remedy according to Me is to pressurise the CPD BEFORE COOLING IT. BTW we heat and cool the CPD by circulating water through the jacket using a waterbath heater/mixer (Lauda type T, -20deg to 100 deg C.) which has a small centrifugal pump on it and which sits in a 2 Litre stainless tank. At the start we fill up the small tank with ice and add enough water to cover the heater/pump on the mixer. Then we start the pump with the heater off and chill the CPD that way. When we want to warm the CPD we toss away the ice water, replace it with tap water -at- 20 deg or so and turn on the heater which warms the waterbath towards 50 deg.
Notwithstanding all of the above, since the CPD only works if all the seals are good, you must keep a couple of seal kits in your lab ready for quick repairs. Failure of the seals is usually discovered after some poor soul has spent weeks on their experiment and days on processing their tissue only to find the CPD leaks. If you have the capacity to quickly replace any of the seals so their experiment survives your reputation will be much enhanced!
Mel Dickson E.M. Unit, University of NSW Sydney, Australia
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} ------------------------------------------------------------------------ Alex wrote, } 1. When a gold coated specimen is viewed in SEM at ~10k or higher, a } fine structure of gold coating becomes visible. Are there optimal } conditions of sputter coating (sputter current, time, distance } target-specimen, argon pressure, other gas than argon) to minimise the } artefact? } } 2. What is Au-Pd target? I thought that it was just an alloy one, but } a supplier of the coater says that using the alloy target is not } enough, that a coater with simultaneous sputtering Au and Pd from two } different targets is required to produce continuous coating.
Dear Alex, 1. Assuming that you are seeing gold, this indicates that your coating is too thick. When a sample is coated it should take on a bluish color. This is most easistly seen on flat areas where there is no sample. Try using a cover slip to set up the conditions. AS to conditions. Don't rush the coating. It should take about a minute to coat your sample. The manuel that came with the sputter coater will give you a starting point.
2. You are right, it is an alloy target. This target should give you a finer grain size, but the secondary electron return is not as good.
Gregory Rudomen Greg-at-UMIC.SUNYSB.EDU 516-444-3126 University Microscopy Imaging Center S.U.N.Y. Stony Brook
Meeting: Tripod Polisher Workshop Dates: June 6 & 7 1997
Topic: This course will cover all aspects of pre-thinning for TEM and will focus on final thinning via Tripod Polishing. Due to the limited class size and the extensive hands-on opportunities, this course is well suited to novices as well as advanced Tripodders. Attendees will also learn the latest techniques available in ion milling and in Plasma Cleaning for TEM samples.
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} } Can anyone please comment on the storage properties of common EM/LM fixatives. } We have some older 1% glut/4% formaldehyde in PO4 buffer. Is there some rule } of thumb that people are using, should we analyze before use, are there some } references we can access? } Thank you in advance for your comments. } } Regards, } Ken Baker } } The aldehydes oxidise to acids - formic or glutaric. The reaction is less at lower pH so is accelerated when you buffer the solution to pH7. Our rule is to buffer the fixative just before we use it and discard any buffered fixative older than a week.
Mel Dickson }
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} 1. When a gold coated specimen is viewed in SEM at ~10k or higher, a } fine structure of gold coating becomes visible. Are there optimal } conditions of sputter coating (sputter current, time, distance } target-specimen, argon pressure, other gas than argon) to minimise the } artefact?
You will probably need to experiment, since the optimum coating will depend to some extent on your specimen - rough specimens will need a coarser, larger grained coating to prevent charging. In addition, you will get a huge amount of advice from all directions, much of it contradictory! so you'll only find out what is right for you by experiment. Anyway, MY advice is:
Argon is generally the best option and you also want to make sure that the argon isn't contaminated with N or O - this will substantially reduce the sputtering rates. If you're Ar supply is good, then this only requires that you flush the system for a short period before sputtering.
Argon pressure/flow rate should be adjusted so that the plasma is just steady - turn up the Ar flow rate until you get a plasma, and then slowly reduce the flow rate (be slow because there will be a significant lag between changing the flow rate and the system stabilising). At some point, the plasma will start to flicker, and then go out. Increase the flow rate to just higher than the point at which the plasma flickers (obviously, this all needs to be setup either without the specimen in the coater, or with a shutter over the specimen).
Lower voltages and shorter times will tend to produce finer and thinner coatings, respectively. Most suppliers will advise approx 1.5 to 1.8 kV, but you can produce some very nice coatings at 600 to 800 V. You should coat for a period just long enough to stop specimen charging.
} 2. What is Au-Pd target? I thought that it was just an alloy one, but } a supplier of the coater says that using the alloy target is not } enough, that a coater with simultaneous sputtering Au and Pd from two } different targets is required to produce continuous coating.
Au/Pd target is an alloy - I'm not aware of commercial coaters that simultaneously work from a pair of targets. Such an arrangement might be more effective but I want some evidence that it was, and I expect it would be more expensive:) Au/Pd alloy targets are effective at producing finer grained coatings. It seems that the Pd provides nuclei for the Au, leading to more, and smaller Au grains rather than the Au grains growing larger as with a pure Au target.
} Of course, it's better to get a FEG and use low voltage, but I want } to do it with a magnetron sputter coater and conventional JSM 6400. } } Thank you, } } Alex
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} We solicit your help in: } (1) Sharing knowledge about similar situations
I used to have a lab 50 metres from a (not so busy) railway line. We could detect the vibration from the trains when they were around a kilometre off.
The problem will vary according to the soil type between your lab and the new road. Ideally the road would be on swampy ground and your lab on a platfrom carved out of granite. That would give good decoupling. But basically, any effective solution in you laboratory will cost several thousands per instrument, as each instrument will need to be relocated on some heavy support (thick steel plate, thicker concrete slab, mass is what you need) with some very flexible mounts under it (air-springs are ideal). It may be more cost effective to route the road further away.
} (2) Pointing us to the best published sources about EM lab design, } particularly in regard to vibration and preferred distance from nearby roads
} (3) Pointing us to any expert EM lab design firms from whom we } might get information
The classic reference work is "Design of the Electron Microscope Laboratory" by Ronald H Alderson 1975. It is Volume 4 in "Practical Methods in Electron Microscopy" Editor Audrey M Glauert. American Elsevier ISBN 0444 1087 6. Was still available two years back whem I bought my second copy to share with the architect for my new laboratory. Pages 68-86 deal with mechanical vibration and decoupling/damping systems } } Mel Dickson } } } }
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Hi I am looking for information about ccd analogous or digital cameras to couple to my optical microscope OLYMPUS BX50. Wanted to know of the fact that resolution must be to use it in reconstruction 3D and digital images procesing. If furthermore know the address e-mail of some providing, them would thank that me facilitate it Thank you very much
Fernando Diego Balducci Laboratory of Electron Microscopy School of Engineery - Bioengineery National University of Entre Rios Argentina Phone: 54 43 975100 Fax : 54 43 975077 e-mail : RNBALDUC-at-ARCRIDE.EDU.AR
1997 ADVANCED SPECIMEN PREPARATION WORKSHOPS (for LM, SEM/TEM, and SPM)
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***Site-specific Cross-sectioning and Microthinning Precision Cleaving (May 6 or Nov. 4 in Santa Clara, CA) FIB-milling for SEM Cross-sectioning (May 7 or Nov.5 in Sunnyvale, CA) FIB-milling for TEM Cross-sectioning (May 7-9 or Nov.5-7 in Sunnycale, CA) Precision Lapping for SEM Cross-sectioning (May 14 or Nov. 12 in Phoenix, AZ) TEM Wedge-polishing (May 14-16 or Nov. 12-14 in Phoenix, AZ)
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Dear Alex, You should not see the structure of a Au-Pd film at 10K. In the 1980 SEM Inc. Conference Proceedings there is an extensive study of the various conditions for making films, with the films studied by TEM. It is worth reading if you have access to it. The result was a recommendation to use Ar gas, 60%Au-40%Pd instead of Au and a lower voltage, 600 to 700 volts. The specimen-surface distance is about 2 cm. I have been using a 1 to 3 minute coating in Ar at 700v. ever since and can only see the film at 50,000 times. Coatings using W or Ni were even finer. I have never heard of using two targets instead one alloyed one. I'm sure the ionized particles don't care. The very fine coatings required by FEG microscopes are best made by an ion-beam coater (much more expensive). You wrote: } Two questions: } } 1. When a gold coated specimen is viewed in SEM at ~10k or higher, a } fine structure of gold coating becomes visible. Are there optimal } conditions of sputter coating (sputter current, time, distance } target-specimen, argon pressure, other gas than argon) to minimise the } artefact? } } 2. What is Au-Pd target? I thought that it was just an alloy one, but } a supplier of the coater says that using the alloy target is not } enough, that a coater with simultaneous sputtering Au and Pd from two } different targets is required to produce continuous coating. } } Of course, it's better to get a FEG and use low voltage, but I want } to do it with a magnetron sputter coater and conventional JSM 6400. } } Thank you, } } Alex Good luck, Mary
I bought a gold/palladium coater recently and found that the default time listed in the manual (} 1 minute) resulted in a coating about the thickness of the chrome on a 1950's Buick. I now coat even the most problematic samples (Mo oxide crystals, glass fractures, rare-earth phosphor particles, etc.) for 15 seconds or less. I also dropped the current to about 70% of the spec value. The samples still generate Au and/or Pd x-rays occasionaly, but I still detect light elements. It's a balancing act.
Remember, it is easier to recoat than de-coat.
my 2 cents.
------------------------------------------------ Opinions or statements expressed herein, rational or otherwise, do not necessarily reflect those of my employer.
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To all on the list I am looking for a lab or person that can do lead phase speciation to distinguish lead products from a smelter from naturally occuring lead minerals. Please reply as soon as possible, there is probably some money and work involved.
Clarissa Vierrether The Doe run Company 573-244-8109 Viburnum, MO
------------------------------------------------------------------------=20 The Microscopy ListServer -- Sponsor: The Microscopy Society of America=20 =20 =20 Alex; =20 In my experience, it would be very unusal to be able to see any detail= s of=20 a Au sputtered coating of normal thickness (5-20 nm.) at a magnificati= on of=20 only 10,000x. Coating artifacts usually become a problem at much high= er=20 magnification such as 50,000x and above. =20 If your sputtered coating is very thick this could be a problem and yo= u=20 should review the operation of your specific coater to determine the b= est=20 parameters for the coating thickness that you desire. The sputter curr= ent,=20 gas pressure, gas quality, distance to the target, and sputtering tim= e are=20 all factors that must be considered. =20 While Au provides an efficient emitter of secondary electrons in the S= EM,=20 the grain size is quite large and does become a problem at higher mags.= =20 Au/Pd is usually an alloy target that combines the better secondary=20 electron emission characteristics of Au along with the smaller grain s= ize=20 of Pd. I've never seen a sputter coater for SEM sample prep that had=20= two=20 targets mounted and ready to go in the same pump down cycle, this soun= ds=20 like a device not designed for SEM sample prep but for industrial sput= ter=20 applications. =20 =20 Pt and Cr are other common target materials that you might also consid= er,=20 however I suspect your problem is not the coating but more likely the=20 sample. You did not state what the sample was so I am guessing that t= here=20 may be some sample deformation of the sample related to vacuum=20 incompatibility or heat from the coater. This could be shrinkage from= the=20 evacuation of the chamber or heat from the magnetron sputter head. =20 If there is a possibility that the sample may be the problem, try sput= ter=20 coating your sample and a piece of metal or carbon disc at the same ti= me. =20 If the coating is the source of the artifact the artifact should be=20 observable on all of the samples. =20 =20 I have had some problems with my server, could you acknowledge receipt= of=20 this post. Thanks and good luck. =20 Hope this helps. =20 John Humenansky Braun Intertec Corp. 6875 Washington Ave. So. Minneapolis, MN 55439 (612) 942-4822 =20 =20 =20
In addition the summary of responses on the Polaron E3000 CPD from Susan Carbyn:
I agree with most of the procedures outlined in your summary and have had frequent leaks with the front seal leaks on our Polaron E3000 since 1992. The unit was put into service in 1979 and from that time until October 1992 I replaced 5 door and 8 Dowtey window seals for 275 drying runs (ave. 34 runs/window seal). Since October 1992 I have used 25 window seals for 113 drying runs (ave. 5 runs/window seal). I am still using the door seal that was installed in March 1990! It has some fine cracks but works every time.
Most recently I have had severe leak problems due to a few defective seals as the manufacturer has mentioned. I am waiting for new replacements. Last month out of desperation for another seal I installed a used one (installed in November 1990) that was removed from the window for preventative maintenance (after 38 runs) in October 1992. It still works ok after 7 more runs.
We use only ethanol, have been heating and cooling with the same system since the unit was installed and always pressurize between 22 and 19 degrees centigrade. I have looked carefully at the surface of the rubber on the newer 1990's seals compared to the used 1980's ones (I inspect all of my seals with a light microscope before installation.). The rubber on the old 80's seals is very smooth and even on the sides and edges. The seals I purchased in the 90's are striated on the sides and have irregular edges. Because of my recent experience successfully reusing my old window seal (as well as the 175 runs on my old door seal) I suspect that the rubber and seal tooling may have changed since the late 80's and could be the cause of the increased failure rate.
The only solution I have for this problem is to increase the budget for replacement seals and switch to using HMDS whenever possible.
Regards,
Jim
James S. Romanow The University of Connecticut Physiology and Neurobiology Department Electron Microscopy Facility U-131 Storrs, CT 06269 bsgphy3-at-uconnvm.uconn.edu 860 486-2914 voice 860 486-1936 fax
I'm posting this for a colleague not on the list. Please respond to Daniel Wilcox at wilcox-at-mailback.macom.com
Daniel is looking for sputter targets for his Technics Hummer 5 sputter coater. (I may not have the manufacturer's name correct, but contact Daniel for exact details.) Any help finding a source is appreciated.
We would like to get super TEM of mouse cardiac muscle. The best technique I know of is to perfuse through the abdominal vena cava, aiming toward the heart, with 4%paraform/2% Glut in cacodylate buffer. Should I use saline first to flush out the blood, should I use pressure, either syringe or gravity? Should I forget the above and listen to some of your comments. I'd appreciate your advice. Thanks so much.
John Hardy City of Hope Medical Center EM Lab jhardy-at-smtplink.coh.org
Hello All, I have been asked recently if there is any way to image micelles, such as those formed by lubricant additives. I thought that light microscopy might work, and possibly confocal. Does anyone have experience in doing this? I have a feeling I will have to refer this work to an outside lab. Regards, Melanie Behrens
Thank you very much All answered the questions about optimal conditions for sputter coating. Reducing Argon pressure and voltage works great even with a gold target!
Thanks again, Alex _________________ Alexander Titkov
Millennium Inorganic Chemicals PO Box 245 Bunbury WA 6231 Australia Ph (097) 808 505 FAX: (097) 808 500 E-mail: scm!atitkov-at-scmaust.attmail.com
At 23:14 9/04/97 -0400, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
These can be imaged using cryo-TEM. Drs Katarina Edwards and Mats Almgren at Dept of Physical Chem, Uppsala University in Sweden have done lots of this type of imaging. I am a bit out of date on this but have some papers.
Gustafsson-J; Arvidson-G; Karlsson-G; Almgren-M, Complexes between cationic liposomes and DNA visualized by cryo-TEM. Biochim-Biophys-Acta. 1995 May 4; 1235(2): 305-12
Edwards et al 1989 Langmuir 5 473-378
and also Edwards and Almgren 1991 Solubilization of lecithin vesicles by C12E8- Structural transitions and temperature effects. J Colloid Interface Sci. I dont have the number for this
Mail address, Mats Almgren, Dept Physical Chemistry, Uppsala University, PO Box 532, S-751 21 Uppsala, Sweden.
this may provide a lead
Dr. Bridget Southwell Department of Anatomy and Cell Biology University of Melbourne Parkville Vic 3052 AUSTRALIA Tel: +61 3 9344-7646 Fax: +61 3 9347-5219
At 23:14 9/04/97 -0400, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
These can be imaged using cryo-TEM. Drs Katarina Edwards and Mats Almgren at Dept of Physical Chem, Uppsala University in Sweden have done lots of this type of imaging. I am a bit out of date on this but have some papers.
Gustafsson-J; Arvidson-G; Karlsson-G; Almgren-M, Complexes between cationic liposomes and DNA visualized by cryo-TEM. Biochim-Biophys-Acta. 1995 May 4; 1235(2): 305-12
Edwards et al 1989 Langmuir 5 473-378
and also Edwards and Almgren 1991 Solubilization of lecithin vesicles by C12E8- Structural transitions and temperature effects. J Colloid Interface Sci. I dont have the number for this
Mail address, Mats Almgren, Dept Physical Chemistry, Uppsala University, PO Box 532, S-751 21 Uppsala, Sweden.
this may provide a lead
Dr. Bridget Southwell Department of Anatomy and Cell Biology University of Melbourne Parkville Vic 3052 AUSTRALIA Tel: +61 3 9344-7646 Fax: +61 3 9347-5219
I am surprised to note the high frequency of door seal leaks experienced by users of the Polaron CPD apparatus. We have two such units which have been in use since the mid-1970's and we have experienced very few of these problems. The procedures we use appear to be similar to those used by the people experiencing these frequent leaks. Perhaps, therefore, Jim Romanow's suggestion that the earlier seals, or even the CPD units themselves, were less prone to failure than the newer ones is correct. Having said that we seldom experience these problems, we did have a chamber door leak a few days ago but this was solved immediately by cleaning the door seat and replacing the seal with a spare which we had. This spare was from a set of spare seals purchased in the early 1980's so was probably of the vintage which Jim suggests has a better record than the more recently-manufactured seals.
Robin H Cross Director : EM Unit, Rhodes University, Grahamstown, South Africa eurc-at-giraffe.ru.ac.za - tel: +27 461 318168 - fax: +27 461 24377
} Hello All, } I have been asked recently if there is any way to image micelles, such as } those formed by lubricant additives. I thought that light microscopy might } work, and possibly confocal. Does anyone have experience in doing this? I } have a feeling I will have to refer this work to an outside lab. } Regards, } Melanie Behrens
I've seen some nice images of this type of specimen produced using cryo-TEM. You might want to check with a lab that is in the food or cosmetics industry.
Regards,
--------------------------------------------------------------- Dr L. P. Stoter Technical Editor, MICROSCOPY & ANALYSIS Technesis 17, Rocks Park Road email: LPS-at-teknesis.demon.co.uk Uckfield, E. Sussex Phone: +44 (0)1825 766911 TN22 2AT Fax: +44 (0)1825 766911 United Kingdom ---------------------------------------------------------------
} I have been asked recently if there is any way to image micelles, such as } those formed by lubricant additives. I thought that light microscopy might } work, and possibly confocal. Does anyone have experience in doing this? I } have a feeling I will have to refer this work to an outside lab.
Bridget Southwell (B.Southwell-at-Anatomy.UniMelb.EDU.AU) suggested the use of CryoTEM and referred to the work of M. Almgren et al. Several groups have successfully applied cryoTEM to imaging micelles. You might want to look at the recent papers from Y. Talmon' group and some older work (1989-1990) by Phillip Vinson.
One caution: most of this work (and my own as well) has studied oil-in-water systems ("normal" micelles.) These systems are relatively easy to image by cryoTEM. I suspect that "lubricant additives" are water-in-oil systems. Several people have attempted cryoTEM on water-in-oil systems (inverse micelles) and have experienced problems with extreme sensitivity to radiation damage. There is some real interesting radiation chemistry that occurs when one irradiates large concentrations of hydrocarbons in the presence of vitreous ice. -- Best Regards, John Minter
Eastman Kodak Company Phone: (716) 722-3407 Analytical Technology Division FAX: (716) 477-3029 Room 2112 Bldg 49 Kodak Park Site email: minter-at-kodak.com Rochester, NY 14562-3712 calendar: via PROFS
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John, The best fixation I've done and ever seen was to simply dice the heart in Millonigs phosphate buffered glutaraldehyde, wash in Millonigs buffer and post fix in Millonigs OsO4. Ian.
We routinely do cardiac perfusions on mice to collect various tissues. We use 2.5% glutaraldehyde in cacodylate buffer and a perfusion pump. Our procedure is as follows.
Set up the perfusion pump with 2 x 30 ml syringes (using a threeway stopcock) and a 21 gauge butterfly needle. Adjust it to deliver a constant rate of perfusate (3 ml/min for 3-6 wk old mice, 6 ml/min for mice 7 wk or older).
With the mouse well anaesthetized, hold the xyphoid process with a hemostat and cut along either side of the sternum along the rib cage allowing the chest wall to be reflected. Insert the needle into the left ventricle and cut the right atrium immediately to allow the perufsate to escape. Wash for 10 min using lactate Ringers or until 30 ml has cleared. Switch to the fixative and fix for 10 min (or 30 ml.) without retracting the needle. Remove any desired tissue samples and continue fixation for 2 h - overnight at 4 degrees C. Then process tissues as usual.
I don't know if the technique would damage the cardiac muscle that you are interested in but I would definitely flush with saline first. As to the syringe/gravity pressure question, we find that while gravity works very well for larger animals such as rats, it's hard to get the right constant pressure on small mice. If you want any more details, let me know.
Pat Hales Dept. of Anatomy & Cell Biology McGill University hales-at-hippo.medcor.mcgill.ca
} Hello All, } I have been asked recently if there is any way to image micelles, such as } those formed by lubricant additives. I thought that light microscopy might } work, and possibly confocal. Does anyone have experience in doing this? I } have a feeling I will have to refer this work to an outside lab. } Regards, } Melanie Behrens
How large? The detergents I am familiar with form micelles no larger than the size of globular proteins, so light microscopy is of little use.
If they are large enough for light microscopy, I would try differential interference contrast (Nomarski), which should yield a good image of the edge of the micelle against the background.
Jim Williams.
/////////////////////////////////////////////////////////////////////////// / James C. Williams, Jr. williams-at-anatomy.iupui.edu / / Department of Anatomy / / Indiana University School of Medicine (317)274-3423 / / 635 Barnhill Drive (317)278-2040 fax / / Indianapolis, IN 46202-5120 / /////////////////////////////////////////////////////////////////////////// Great are the works of the LORD, studied by all who have pleasure in them. Psalm 111:2
} We would like to get super TEM of mouse cardiac muscle. The best } technique I know of is to perfuse through the abdominal vena cava, } aiming toward the heart, with 4%paraform/2% Glut in cacodylate buffer. } Should I use saline first to flush out the blood, should I use } pressure, either syringe or gravity? Should I forget the above and } listen to some of your comments. I'd appreciate your advice. Thanks } so much. } } John Hardy } City of Hope Medical Center } EM Lab } jhardy-at-smtplink.coh.org
John- Definitely flush first, otherwise the blood will coagulate in the fix and block the flow of blood to the heart. You don't say what animal you are using, but in general any vessel close to and with as direct as possible access to the vessels of interest can be used for perfusion. I have had good luck in perfusing the coronary arteries and thus also the heart muscle in rats by inserting the perfusion needle directly into the left ventricle. This has the added advantage of distributing fixative to the inner wall of the heart. We inject with heparin just before anesthesia as an added measure against coagulation during manipulation, cannulation, and before flushing of the artery is complete. I prefer to flush with the same buffer used for the fixative (cacodylate) rather than saline. I don't have imperical data, however, to prove one type of flush is better than another. We often do enzyme or immunohistochemistry on tissue and my protocols are often based on the "if it ain't broke don't fix it" model and my belief that the best histochemist are really voodoo priests in disguise.
I hope this helps Jay ************************************************************** * aka: W. Gray Jerome * * Dept. of Pathology * * Bowman Gray School of Medicine of Wake Forest University * * Medical Center Blvd * * Winston-Salem, NC 27157-1092 * * 910-716-4972 * * jjerome-at-bgsm.edu * **************************************************************
Thank you to everyone who has sent information about Hummer targets. Daniel Wilcox said that his network has been experiencing problems, so I have been forwarding messages sent to me. My e-mail address is audrey.dow-at-amp.com
In response to the following: ============================================== I have been asked recently if there is any way to image micelles, such as those formed by lubricant additives. I thought that light microscopy might work, and possibly confocal. Does anyone have experience in doing this? I have a feeling I will have to refer this work to an outside lab. ============================================== One of the earliest pieces of work I can remember being published came out of the old Sinclair Research Laboratories on the South Side of Chicago. I regret I can not remember names, because some of their work was published (some not) in the early 1960's but they had beautiful images of features from lubricants, having the appearance sometimes of almost filamentous like structures. These were before the days of SEM and hence, the work was all done by Pt/C replication techniques and TEM. Note: Even if SEM was around, the dimensions of the structures were such that the features would have been difficult to impossible to resolve anyhow. Off line, if anyone is interested, I would related more on the history of how the technique was developed, which was also influenced by Mr. Bill Ladd, founder of Ladd Reserach Industries, Inc.
So if the Pt/C replication approach will show what you are looking for, it will be faster and cheaper than any other method. Now I am not sure that these filamentous structures would qualify strictly speaking as "micelles", but more often than not, in our own lab, when our clients ask to see the "micellar structure", in these kinds of materials, this is what they often times mean, because they pull out some old micrographs showing the filamentous structures! In any case, we have been practicing this technique ever since the early 1970's on lublicant samples of various types.
If what you want to see is more along the lines of a traditional micellar structure, then you have to turn to freeze fracture TEM, however, the nature of the organics present, and the need to clean off the organics (often times polymers found in today's lubricant systems) that "stick" to the replica make this approach often times a lot more tricky than might originally meet the eye. It can be, in others words, an art unto itself. But structures can be seen, they tend to be (at least the ones we have seen) an almost "network" or "fishnet" kind of structure. Maybe other structures are present as well, but these are the kinds of structures we have seen ourselves.
Disclaimer: Our firm, Structure Probe, Inc. performs these kinds of analyses as an analytical laboratory service.
Chuck
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--------------------------------------------------------------- Dr L. P. Stoter Technical Editor, MICROSCOPY & ANALYSIS Technesis 17, Rocks Park Road email: LPS-at-teknesis.demon.co.uk Uckfield, E. Sussex Phone: +44 (0)1825 766911 TN22 2AT Fax: +44 (0)1825 766911 United Kingdom ---------------------------------------------------------------
} Hello All, } I have been asked recently if there is any way to image micelles, such as } those formed by lubricant additives. I thought that light microscopy might } work, and possibly confocal. Does anyone have experience in doing this? I } have a feeling I will have to refer this work to an outside lab. } Regards, } Melanie Behrens
Cryotechnique is the way to go, but "cryo-TEM" isn't the best approach. I recommend freeze-fracture; see Robards & Sletyr, "Low temperature methods in biological EM" (v. 10 of the Glauert series), 1985, and Zazadzinski & Bailey,"Freeze-fracture of polymers", J.E.M. Technique 13:309-334(1989). There are commercial labs with the necessary equipment.
Caroline Schooley Educational Outreach Coordinator Microscopy Society of America Box 117, 45301 Caspar Point Road Caspar, CA 95420 Phone/FAX (707)964-9460 http://www.MSA.microscopy.com/ProjectMICRO/Books.html
I am new to this list and am trying to do some immunogold labeling studies (I am also a novice in this area). I have a problem that I can't figure out and thought someone out there could help me. It is probably a simple one, but so far I can not find a solution. I am working with samples that are pretty fragile and found that I need to place them on coated grids for stability. I am coating my nickel grids with Pioloform and have found that my sample looks good on these grids. I tried to immunogold label and found non specific labeling throughout the sample, on the resin, and on the grid itself. I have now done tests on the coated grid alone and have processesd it as I would normally. I find that I have gold label everywhere on the coated grid. Therefore I have concluded that an interaction between my primary antibody and the coating is occuring (I have done a control experiment leaving out the primary antibody in the processing and know nothing binds). What could cause this? My blocking solution consists of 1% BSA in TBS +.05% Tween 20 + .02% azide for preservation. Could it be that the blocking solution is not doing its job? How long can you store blocking solution? Could I be using a too concentrated antibody solution? Could this be specific to my antibody? I should mention that I wash my grids by either passing them through drops of TBS or wash using a light stream of TBS from a pasteur pipet. Both methods yield the same results. If anyone can give me some answers to my questions I would appreciate it. Thanks for your time.
I'm looking for suppliers of Ellis type fiberoptic light scramblers. I have found one company, Technical Video in Woods Hole. Are there others you can recommend?
We routinely do cardiac perfusions on mice to collect various tissues. We use 2.5% glutaraldehyde in cacodylate buffer and a perfusion pump. Our procedure is as follows.
Set up the perfusion pump with 2 x 30 ml syringes (using a threeway stopcock) and a 21 gauge butterfly needle. Adjust it to deliver a constant rate of perfusate (3 ml/min for 3-6 wk old mice, 6 ml/min for mice 7 wk or older).
With the mouse well anaesthetized, hold the xyphoid process with a hemostat and cut along either side of the sternum along the rib cage allowing the chest wall to be reflected. Insert the needle into the left ventricle and cut the right atrium immediately to allow the perufsate to escape. Wash for 10 min using lactate Ringers or until 30 ml has cleared. Switch to the fixative and fix for 10 min (or 30 ml.) without retracting the needle. Remove any desired tissue samples and continue fixation for 2 h - overnight at 4 degrees C. Then process tissues as usual.
I don't know if the technique would damage the cardiac muscle that you are interested in but I would definitely flush with saline first. As to the syringe/gravity pressure question, we find that while gravity works very well for larger animals such as rats, it's hard to get the right constant pressure on small mice. If you want any more details, let me know.
Pat Hales Dept. of Anatomy & Cell Biology McGill University hales-at-hippo.medcor.mcgill.ca
Caroline Schooley recommended that Melanie Behrens use freeze fracture to image micelles in her lubricant's. Our lab did some comparisons between freeze-fracture TEM and direct imaging of vitrified micelles of cetyltrimethylammonium chloride (CTACl)/ sodium salicylate (NaSal). Both the CTACl globular micelles and worm-like micelles formed with the addition of NaSal to CTACl are easily imaged by direct cryoTEM. We were unable to obtain contrast in freeze-fracture images without etching. The structures observed in freeze-fracture/freeze-etch micrographs of these systems were much larger than those observed by direct cryoTEM. ALWAYS beware of changes upon etching these microstructures. Our conclusions were that the freeze-fracture, freeze-etch micrographs were dominated by artifacts. I'd be interested in hearing from anyone who had obtained convincing evidence of micellar microstructures by freeze-frature.
-- Best Regards, John Minter
Eastman Kodak Company Phone: (716) 722-3407 Analytical Technology Division FAX: (716) 477-3029 Room 2112 Bldg 49 Kodak Park Site email: minter-at-kodak.com Rochester, NY 14562-3712 calendar: via PROFS
Hello microscopists, I am in the process of assembling an ultra high vacuum chamber with mag. lev. turbo pumps backed by oil free diaphragm pumps. Some parts e.g. gauges on this system will come from other decommissioned systems. I am wondering if someone would like to share experience in: 1. converting viton onto metal O-ring sealings as compared to redesigning flanges to accomodate KFs; including costs, companies, and suppliers; 2. reliability, service record, and major troubles with Leybold, Balzers/Pfeiffer, Osaka, etc magnetic bearing turbo pumps. Sincerely, Marek Malecki.
This should be a simple procedure but I am having a terrible time trying to stain some Embed 812 embedded tissues with uranyl acetate/lead citrate. The tissue is from frog tadpoles, fixed in glutaraldehye/paraformaldehyde and postfixed in osmium. I have tried staining for 5-20 minutes with saturated solution of UA in ethanol, followed by 1-5 minutes in lead citrate (Venable-Coggeshall formulation). The sections look no different from unstained specimens.
I'd appreciate any suggestions about what I might be doing wrong.
Gary Radice, Associate Professor gradice-at-richmond.edu Department of Biology 804-289-8107 (voice) University of Richmond VA 23173 804-289-8233 (FAX)
Leitz has built one several years ago (called Variolum) for their Diaplan, Aristoplan microscopes. I have one of these. Although intended for the regulation of light intensity of xenon and Hg arcs it performs well in video microscopy. You should contact Leitz, or some part dealers to find one.
Does anyone have a good protocol for embedding tadpole brain for doing immunocytochemistry? We have been trying and seem to be having a problem with infiltration. Please let me know. Thanks in advance.
Mei Lie Wong Department of Biochemistry HHMI-UCSF Ph. 415-476-4441 Fax 415-476-1902 email wong-at-msg.ucsf.edu
Dear Susan, You do not say in your message what the antibodies are directed against. One very obvious possibility is that there is specific reactivity to BSA. This would, of course, result in very specific binding to the BSA covering the sample, the resin and the film. This would look as if there was non-specific binding.
A simple test is to change your blocking agent. Try 1% cold-water fish skin gelatin (very cheap, from Sigma) in PBS as a substitute. It is useful because there are no mamalian serum proteins present and for anyone interested in localizing biotin with antibodies, there are no biotin-like molecules present.
If changing the blocking agent doesn't work, I would suspect that the antibody specifically reacts with the resin and the plastic (just kidding!).
There should be no non-specific reactions between the antibodies and the plastic film or resin. Suspect instead the blocking agent or as a last resort, the antibody.
I have included a section on reasons why antibody labeling may not work on my web site.
regards,
Paul Webster, Ph.D. Center for Cell Imaging Yale School of Medicine http://info.med.yale.edu/cellimg --------------------------------------
Hello,
I am new to this list and am trying to do some immunogold labeling studies (I am also a novice in this area). I have a problem that I can't figure out and thought someone out there could help me. It is probably a simple one, but so far I can not find a solution. I am working with samples that are pretty fragile and found that I need to place them on coated grids for stability. I am coating my nickel grids with Pioloform and have found that my sample looks good on these grids. I tried to immunogold label and found non specific labeling throughout the sample, on the resin, and on the grid itself. I have now done tests on the coated grid alone and have processesd it as I would normally. I find that I have gold label everywhere on the coated grid. Therefore I have concluded that an interaction between my primary antibody and the coating is occuring (I have done a control experiment leaving out the primary antibody in the processing and know nothing binds). What could cause this? My blocking solution consists of 1% BSA in TBS +.05% Tween 20 + .02% azide for preservation. Could it be that the blocking solution is not doing its job? How long can you store blocking solution? Could I be using a too concentrated antibody solution? Could this be specific to my antibody? I should mention that I wash my grids by either passing them through drops of TBS or wash using a light stream of TBS from a pasteur pipet. Both methods yield the same results. If anyone can give me some answers to my questions I would appreciate it. Thanks for your time.
Susan
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Hello!
We have had several things to cause this in our sections:
1. Specimen was not osmicated enough, OsO4 too dilute, or bad.
2. or... The lead stain was not the right ph and actually bleached the sample { goal~ = 12 pH} ( we made a switch in Lead stain types just 2 months ago for this very reason.)
3. or.... Some epoxy mixtures, if not polymerized enough or infiltrated ( acts like a really soft block) then the thicks will overstain and the thins will understain. In doing some epoxy mixture experiments, I found with the same chemicals, that certain ratios caused some more hard to stain sections.
4. or..... someone made a mistake by a factor of 10 in the NaOH [ ] of 1st rinse dip after lead stain, and it bleached the dickens out of the section. We had a researcher do this to their sections.
5. or..... we have bad water or something in our building, our aquous Uranyl Acetate only does well for 10 days, so we make it up in very very small amounts. I like the clearer background we get with aqueous much better than the alcohol methods, but that is personal preference.
6. I've developed a microwave technique, and my grids stain really well now. Write if you want to hear more....it's too lengthy for this message.
We stopped using Epon812 about 7 years ago, because at the time it had a lot of impurities in it, the resin ate up our diamond knives. And the blocks were real difficult, poor polymerization despite how we would polymerize it. Switched to Lx112 from Ladd and our diamonds last 6 X longer.
Hope some of this helps, Lou Ann
} This should be a simple procedure but I am having a terrible time trying to } stain some Embed 812 embedded tissues with uranyl acetate/lead citrate. The } tissue is from frog tadpoles, fixed in glutaraldehye/paraformaldehyde and } postfixed in osmium. I have tried staining for 5-20 minutes with saturated } solution of UA in ethanol, followed by 1-5 minutes in lead citrate } (Venable-Coggeshall formulation). The sections look no different from } unstained specimens. } } I'd appreciate any suggestions about what I might be doing wrong. } } } Gary Radice, Associate Professor gradice-at-richmond.edu } Department of Biology 804-289-8107 (voice) } University of Richmond VA 23173 804-289-8233 (FAX)
Lou Ann Miller Center for Microscopy & Imaging College of Veterinary Medicine Dept. of Veterinary Biosciences University of Illinois Rm 1108 Basic Sciences Bld 2001 S Lincoln Ave. Urbana, Illinois 61802
Phone: 217-244-1567 email: lamiller-at-uiuc.edu
Center for Microscopy & Imaging Home page: http://www.cvm.uiuc.edu/MicImagLab/MicImagLab.html
Central States Microscopy Society: http://www.cvm.uiuc.edu/Homepages/LouAnnMiller/CSMS/csms.html
Personal Home Pages: http://www.cvm.uiuc.edu/Homepages/LouAnnMiller/LAM.html
Attention ESEM owners and users: If you were desperate for the low magnification in ESEM (E3, 2020) in a HiVac mode do not worry from now on. Simply, take out the rotation/tilt module and put your specimen directly on the driving assembly. You will be deprived of rotation and tilting of a specimen but you will gain extra 62 mm of working distance. Alternatively, put you sample in a brass or aluminium cylinder with a side screw to keep the specimen firm and glue the cylinder on the driving assembly with a double-sided conductive tape. The height of cylinder is such that it can accomodate a full lenght of a pin of a pin-type stub. In this case you will gain 'only' about 50 mm of working distance. Even without playing with apertures you should get minimum mag ~6X at acc. voltage of 2 kV, covering an area of 13 mm in diameter with a much better collecting angle for SE, when using ET detector. Try for yourself and enjoy the results. Cheers, Wis Jablonski, OiC EM/X-Ray Microanalysis, Uni of Tasmania
Hello! A student at our department shall try to label membrane proteins on mouse spermatozoa for study with SEM. We have access to cryo SEM as well which might be the fastest procedure. Conventional technique with chemical fixation and labelling of single cells or label fresh (living) spermatozoa and then use cryotechniques? Any hints will be appreciated.
Hans Ekwall Centre for Reproductive Biology Dept. of Anatomy & Histology SLU, Box 7011, 75007 Uppsala SWEDEN
Has anyone comments on the UV polimeriztion of Lowicryl HM20 after freeze substitution in Acetone-Uranylacetate. I have problems with prepolymerization of the resin which causes unusable blocks, but the problem is not consistent, some blocks are good some are not.
TIA, Stefan
Dr. Stefan Hillmer Albrecht-von-Haller Institut fuer Pflanzenwissenschaften Universitaet Goettingen Untere Karspuele 2 37073 Goettingen Deutschland
Tel (+49) 551-392013 Fax (+49) 551-397833 e-mail shillme-at-gwdg.de
we have had similar problems on pioloform coated grids, and I would say that your 1. AB is binding to the pioloform. Solution: Try change the coating to formvar or a carbon coating. You can also make the grids float on the 1. AB so that the ABs only get in contact with the pioloform outside the sections.
Bo
} Hello, } } I am new to this list and am trying to do some immunogold labeling } studies (I am also a novice in this area). I have a problem that I } can't figure out and thought someone out there could help me. It is } probably a simple one, but so far I can not find a solution. I am } working with samples that are pretty fragile and found that I need to } place them on coated grids for stability. I am coating my nickel grids } with Pioloform and have found that my sample looks good on these grids. } I tried to immunogold label and found non specific labeling throughout } the sample, on the resin, and on the grid itself. I have now done tests } on the coated grid alone and have processesd it as I would normally. I } find that I have gold label everywhere on the coated grid. Therefore I } have concluded that an interaction between my primary antibody and the } coating is occuring (I have done a control experiment leaving out the } primary antibody in the processing and know nothing binds). What could } cause this? My blocking solution consists of 1% BSA in TBS +.05% Tween } 20 + .02% azide for preservation. Could it be that the blocking solution } is not doing its job? How long can you store blocking solution? Could I } be using a too concentrated antibody solution? Could this be specific to } my antibody? I should mention that I wash my grids by either passing } them through drops of TBS or wash using a light stream of TBS from a } pasteur pipet. Both methods yield the same results. If anyone can give } me some answers to my questions I would appreciate it. Thanks for your } time. } } Susan } }
Advanced International Immunofluorescence Course Gargnano '97 (Italy)
The Advanced International Immunofluorescence Course is a post-doctorate theoretical/practical course, with propedeutical lectures and practical stages on traditional and confocal immunofluorescence microscopy and image and ion analysis. The course will take place in Gargnano (Lake of Garda) from 7 to 10 October 1997. Further information and registration details will be found at the following Web address
http://imiucca.csi.unimi.it/endomi/ACIF.html
Thank you Paolo Castano ____________________________________________________________________________ _______
Prof. Paolo Castano UNIVERSITY OF MILAN INSTITUTE OF HUMAN ANATOMY - CHAIR OF HUMAN ANATOMY FOR PHARMACY Via Mangiagalli, 31 - 20133 Milan (Italy)
} I have been asked recently if there is any way to image micelles, such as } those formed by lubricant additives. I thought that light microscopy might } work, and possibly confocal. Does anyone have experience in doing this? I } have a feeling I will have to refer this work to an outside lab.
Bridget Southwell (B.Southwell-at-Anatomy.UniMelb.EDU.AU) suggested the use of CryoTEM and referred to the work of M. Almgren et al. Several groups have successfully applied cryoTEM to imaging micelles. You might want to look at the recent papers from Y. Talmon' group and some older work (1989-1990) by Phillip Vinson.
One caution: most of this work (and my own as well) has studied oil-in-water systems ("normal" micelles.) These systems are relatively easy to image by cryoTEM. I suspect that "lubricant additives" are water-in-oil systems. Several people have attempted cryoTEM on water-in-oil systems (inverse micelles) and have experienced problems with extreme sensitivity to radiation damage. There is some real interesting radiation chemistry that occurs when one irradiates large concentrations of hydrocarbons in the presence of vitreous ice. -- Best Regards, John Minter
Eastman Kodak Company Phone: (716) 722-3407 Analytical Technology Division FAX: (716) 477-3029 Room 2112 Bldg 49 Kodak Park Site email: minter-at-kodak.com Rochester, NY 14562-3712 calendar: via PROFS
Does anyone have any thoughts or experience with using a modern (e.g. Generation 3) image intensifier tube to enhance CCD imaging on a TEM? I don't know the limitations of image intensifiers (e.g. noise, resolution) but the potential for increased gain on high-resolution, short exposure time shots might be worth investigating.
Daniel L. Callahan Mechanical Engineering and Materials Science Rice University, MS 321 6100 S. Main St Houston, TX 77005-1892
Thanks to all who have responded to the call for lead speciation in smelter vs natural occuring products. The information has been forwarded to the correct person and they will make there decision soon because the final report is due in 5 weeks. Thanks again.
Clarissa Vierrether The Doe Run Co. cvierret-at-misn.com
I have never tried Pioloform but routinly use either parlodion coated nickel or formvar coated nickel grids without backround problems. That would be the first thing I would test. The other possiblity is that the original antigen was bound to BSA as a stablizer when they made the antibody and now the primary is binding to the BSA. Try using purified gelatin instead of BSA for blocking.
On Thu, 10 Apr 1997, Susan Fujimoto wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Hello, } } I am new to this list and am trying to do some immunogold labeling } studies (I am also a novice in this area). I have a problem that I } can't figure out and thought someone out there could help me. It is } probably a simple one, but so far I can not find a solution. I am } working with samples that are pretty fragile and found that I need to } place them on coated grids for stability. I am coating my nickel grids } with Pioloform and have found that my sample looks good on these grids. } I tried to immunogold label and found non specific labeling throughout } the sample, on the resin, and on the grid itself. I have now done tests } on the coated grid alone and have processesd it as I would normally. I } find that I have gold label everywhere on the coated grid. Therefore I } have concluded that an interaction between my primary antibody and the } coating is occuring (I have done a control experiment leaving out the } primary antibody in the processing and know nothing binds). What could } cause this? My blocking solution consists of 1% BSA in TBS +.05% Tween } 20 + .02% azide for preservation. Could it be that the blocking solution } is not doing its job? How long can you store blocking solution? Could I } be using a too concentrated antibody solution? Could this be specific to } my antibody? I should mention that I wash my grids by either passing } them through drops of TBS or wash using a light stream of TBS from a } pasteur pipet. Both methods yield the same results. If anyone can give } me some answers to my questions I would appreciate it. Thanks for your } time. } } Susan }
SHORT COURSE ANNOUNCEMENT ------------------------------------------------- FUNDAMENTALS AND APPLICATIONS OF LIGHT MICROSCOPY ------------------------------------------------- JUNE 22-27, 1997, Burlington Vermont
Experienced microscopy problem solvers will teach a 5-day course on achieving the maximum information from light microscopy. The emphasis of the course will be to provide hands-on experience, and the background for interpretation of images.
The course will cover the principals of light microscopy, contrast techniques for the microscope, adjustments of the microscope for optimum contrast and resolution, interpretation of images in terms of light-matter interactions, and image recording.
A full range of reflected and transmitted light microscopes, as well as contrast accessories, will be provided for use by the students. Students are encouraged to bring their own samples.
Faculty: Philip C. Robinson, author of the RMS Microscopy Series book, "Applied Polarized Light Microscopy", Robert Janes, of the Metropolitan Forensic Science Laboratory, and Mary. McCann, McCann Imaging, course organizer. Vermont Optecs, specialists in research instruments, will supply microscope equipment for the course.
For course brochure and registration information, contact Mary McCann, McCann Imaging e-mail: mccanns-at-tiac.net telephone 617-484-7865 fax: 617-484-7865 Further information: www.microscopyed.com
The french company ALTO MEDIA is producing a series of short movies (3 '), based on a 'travel' into the matter... 5 materials have already been investigated : stell, aluminium, alumina, concrete and brass. These movies require that sequences of both SEM and TEM images are realized. They will be distributed in France (Cite des Sciences, TV broadcasts), and could also be distributed in other european countries (Italy, U.K.,...). ALTO MEDIA is looking for partners, i.e. microscopists, who could be interested to collaborate to this project. The following (non-exhaustive) list gives some ideas of possible interesting materials :
paper/wood/porcelain/plaster/skin/bone/ stone/diamond/graphite/plastics/silicom/iron(rust?)ice/composites(kevlar,mylar)/ ice/glues/lubricating oils,..., and all kinds of materials that are currently used, and for which a microscopic observation can help to understand their properties.
Contact : ALTO MEDIA Etienne Blanchon or Gabriel Turquier tel: 33 01 42 77 77 72 Fax : 33 01 42 77 77 73 e-mail : altomail-at-worlnet. fr
--------------------------------------------------------------------------- Dr. Thierry EPICIER, GEMPPM, umr CNRS 5510, INSA de LYON, Bat 502, 20, Av. Einstein, F69621 VILLEURBANNE CEDEX FRANCE
Hi Gary, We use Embed 812 to process retina tissue. We get intense staining using 4% Uranyl Acetate in 100% methanol.Stain for 30 or 40 minutes at room temp. Keep solution in dark.Rinse in 3 changes of methanol. Then counter stain with Reynold's Lead Citrate for 1-10 minutes. Rinse with 3 changes of DI water. We get beautiful stain if the tissue is well fixed.
Sally
On Thu, 10 Apr 1997, Gary Radice wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } This should be a simple procedure but I am having a terrible time trying to } stain some Embed 812 embedded tissues with uranyl acetate/lead citrate. The } tissue is from frog tadpoles, fixed in glutaraldehye/paraformaldehyde and } postfixed in osmium. I have tried staining for 5-20 minutes with saturated } solution of UA in ethanol, followed by 1-5 minutes in lead citrate } (Venable-Coggeshall formulation). The sections look no different from } unstained specimens. } } I'd appreciate any suggestions about what I might be doing wrong. } } } Gary Radice, Associate Professor gradice-at-richmond.edu } Department of Biology 804-289-8107 (voice) } University of Richmond VA 23173 804-289-8233 (FAX) } } }
Check the following things regarding your problem with prepolymerization of Lowicryl HM20:
1. Are all components being well mixed? The best way to mix the components is to gently bubble nitrogen gas through a glass pipette tip for about 15-20 minutes, or until the benzoyl peroxide catalyst is completely dissolved.
2. Is UV light reaching all sides of the capsules? If you are using polyethylene capsules, are they suspended in wire loops so they are not irradiated from just the tops or the sides?
3. Is the proper UV light being used? Use long-wavelength (366 nm) UV. Short wavelength UV can be too energetic.
4. I would suspect the UV light is too intense. Is the UV light being properly diffused, so it is indirect? Along with this, is the UV source far enough away from the capsules? This depends on the intensity of the UV source, but the UV light should probably be 25-30 cm above the capsules. Also, is there a reflector between the UV source and the capsules, and are all of the inner surfaces of your polymerization chamber lined with aluminum foil? Try a set-up like the following (in cross-section):
O ---------UV source /\ / \ ---------Reflector
uuu ---------Capsules
The easiest way to decrease the intensity of the UV light is to increase the distance between the bulb and the capsules. If you can't do this because of the dimensions inside the freezer or cold box, you can put a layer or two of thin, "frosted" glass in front of the UV light.
I hope this helps. Let us know how things work out.
Best regards, Bob Chiovetti (RCHIOVETTI-at-aol.com)
The staining with heavy metals (Pb, UA) is dependent on proper processing protocols to some extent: Osmium acts as a mordant for UA, and UA acts as a mordant for Pb. Sections which have been exposed to the electron beam are so highly crosslinked due to the high temperature achieved that they may never stain. (Blocks which are overpolymerized in a microwave oven will NEVER stain, no matter what one does) The most likely problem is your Pb stain and your rinsing afterwards. If the pH of the lead stain is over 12 (pH meters are not accurate at these high readings, so one cannot depend on them) there will be NO staining. NEVER, NEVER, NEVER, rinse sections which have just left the lead stain in ANY type of sodium hydroxide solution. This may (will) change the pH radically and erase any stain you may have, or it may cause your stain to "dump". Use plain water. MAKE STAIN CORRECTLY. Use the Reynolds method for lead citrate. NEVER, NEVER, NEVER, use sodium hydroxide pellets to produce the stain. Use commercially titrated sodium hydroxide ( 1.N). Pellets will NOT give you an accurate pH. (and your pH meter is no help, because it functions poorly with this unbuffered solution). Shake and invert the solution for 25 minutes. Boring? Yes! Do it anyway. It is necessary for correct chealation. A lot of information like this is to be found in my chapter in a textbook that came out last year. If you continue to have trouble, please E-mail me, and I will send you a copy of the chapter. ( hcrowley-at-DU.edu ) This type of problem makes one rabid. It drove me crazy. I finally did 4 years of work and observations until I got to the bottom of it. I found that 99% of the time it was the Pb that was so difficult to understand. Good luck. If things do not straighten up, let me know. I would be happy to help you - I know how screaming frustrating it is to have sections which do not stain. Bye, H.
The staining with heavy metals (Pb, UA) is dependent on proper processing protocols to some extent: Osmium acts as a mordant for UA, and UA acts as a mordant for Pb. Sections which have been exposed to the electron beam are so highly crosslinked due to the high temperature achieved that they may never stain. (Blocks which are overpolymerized in a microwave oven will NEVER stain, no matter what one does) The most likely problem is your Pb stain and your rinsing afterwards. If the pH of the lead stain is over 12 (pH meters are not accurate at these high readings, so one cannot depend on them) there will be NO staining. NEVER, NEVER, NEVER, rinse sections which have just left the lead stain in ANY type of sodium hydroxide solution. This may (will) change the pH radically and erase any stain you may have, or it may cause your stain to "dump". Use plain water. MAKE STAIN CORRECTLY. Use the Reynolds method for lead citrate. NEVER, NEVER, NEVER, use sodium hydroxide pellets to produce the stain. Use commercially titrated sodium hydroxide ( 1.N). Pellets will NOT give you an accurate pH. (and your pH meter is no help, because it functions poorly with this unbuffered solution). Shake and invert the solution for 25 minutes. Boring? Yes! Do it anyway. It is necessary for correct chealation. A lot of information like this is to be found in my chapter in a textbook that came out last year. If you continue to have trouble, please E-mail me, and I will send you a copy of the chapter. ( hcrowley-at-DU.edu ) This type of problem makes one rabid. It drove me crazy. I finally did 4 years of work and observations until I got to the bottom of it. I found that 99% of the time it was the Pb that was so difficult to understand. Good luck. If things do not straighten up, let me know. I would be happy to help you - I know how screaming frustrating it is to have sections which do not stain. Bye, H.
Thanks to everyone who offered suggestions for improving my UA/Pb staining of Embed 812 embedded tissues. I now have more staining protocols than I have specimens!
Among the suggestions were
en bloc stain with UA before embedding make up UA in methanol make sure the UA is really saturated by sonicating it make sure tissue is well osmicated check Pb pH is } 12 Try different lead formulation try different epoxy component proportions change to different embedding medium Increase stain time to 30 min (UA) and 5 min(Pb) Try Pb then UA then Pb Try permanganate instead of UA reduce wash times try a microwave stain protocol
Thanks to:
Lou Ann Miller Phil Oshel Robin Cross Julian Smith Tamara Howard David Patton
Gary Radice, Associate Professor gradice-at-richmond.edu Department of Biology 804-289-8107 (voice) University of Richmond VA 23173 804-289-8233 (FAX)
Wis and other ESEM users It is true that longer working distances and lower accelerating voltages in the ESEM give lower mag images and as you point out this is mainly usable at hivac. This is because in wet mode the scattering effect of the gas on the beam is much worse for both longer working distance and lower accelerating voltage dramatically lowering the image contrast to the point that it may not be useable. If the sample is compatible with hivac and you have the luxury of access to other SEMs why not use a conventional SEM? Also at hivac the final pressure limiting aperture in the ESEM can be enlarged such that it does not restrict the scanned beam at low magnification. Further Gene Taylor has come up with a low magnification device which is described in our paper: M.E. Taylor and S.A. Wight "A New Method for Low-Magnification in the Environmental Scanning Electron Microscope" SCANNING Vol. 18, 483-489 (1996). This paper discusses and compares several approaches for attaining low mag. Please see figures 3 and 4 for an explanation and demonstration of the effect of working distance on wet mode imaging.
Scott Wight
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
------------------------------------------------------------------ Scott Wight e-mail: SCOTT.WIGHT-at-NIST.GOV NIST - Microanalysis Group W voice: 301-975-3949 Bld 222, Rm A113 | fax:301-216-1134/301-417-1321 Gaithersburg, MD 20899 \|/ disclaimer: Any opinion expressed is my own and does not represent those of my employer.
My maglev turbo experience is limited to the Leybold 340 l/s pump. I do not
recall when, exactly, I put this pump on my system, but it was shortly after
they became available (} 5 years?). This pump has been running 24 hours/day since, experiencing about 10-15 short time shutdowns from causes like power outages. Shutdown/start-up (according to Leybold - I believe 'em) is harder on the pump than continuous operation. Although it was expensive, it has performed well with zero down-time from failure. I did like the Leybold feature of not needing batteries for "spin-down". During shutdown, it uses the rotor/motor for a generator to supply magnetic bearing power and, thus, apply dynamic braking. BTW....I cannot detect any induced vibration on my rather vibration sensitive Etec SEM.
No vested interest in any pump manufacturer....
Woody White ______________________________ Reply Separator _________________________________
------------------------------------------------------------------------ The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Hello microscopists, I am in the process of assembling an ultra high vacuum chamber with mag. lev. turbo pumps backed by oil free diaphragm pumps. Some parts e.g. gauges on this system will come from other decommissioned systems. I am wondering if someone would like to share experience in: 1. converting viton onto metal O-ring sealings as compared to redesigning flanges to accomodate KFs; including costs, companies, and suppliers; 2. reliability, service record, and major troubles with Leybold, Balzers/Pfeiffer, Osaka, etc magnetic bearing turbo pumps. Sincerely, Marek Malecki.
2:00 5:00 Image analysis workshop, demonstration. Dr. Charlie Meshul's lab. V.A. Bldg. 100, Room 2C-150.
5:15 6:30 Social at Center for Research on Occupational and Environmental Toxicology / Basic Science Building's Atrium.
A quick note for those of you not familiar with our society's meeting format. There is no registration fee. Your annual dues cover registration and also the Friday night social. The social will feature cheeses, cold cuts, breads and fresh fruit and to wash it all down we have an outstanding selection of Northwest wines chosen personally by Charlie Meshul,. Make it a point to attend.
If you have any questions give Bob Mixon, Charlie Meshul, or me a call. There is parking at the VA as noted on the map.
There are tables available for any vendors that attend. If any vendor wants me to put out their literature or samples I am more than happy to help, just let one of the officers know.
Thanks to everyone who offered suggestions for improving my UA/Pb staining of Embed 812 embedded tissues. I now have more staining protocols than I have specimens!
Among the suggestions were
en bloc stain with UA before embedding make up UA in methanol make sure the UA is really saturated by sonicating it make sure tissue is well osmicated check Pb pH is } 12 Try different lead formulation try different epoxy component proportions change to different embedding medium Increase stain time to 30 min (UA) and 5 min(Pb) Try Pb then UA then Pb Try permanganate instead of UA reduce wash times try a microwave stain protocol
Thanks to:
Lou Ann Miller Phil Oshel Robin Cross Julian Smith Tamara Howard David Patton Sally Shrom Krystyna Rybicka
Gary Radice, Associate Professor gradice-at-richmond.edu Department of Biology 804-289-8107 (voice) University of Richmond VA 23173 804-289-8233 (FAX)
Does anyone have any thoughts or experience with using a modern (e.g. Generation 3) image intensifier tube to enhance CCD imaging on a TEM? I don't know the limitations of image intensifiers (e.g. noise, resolution) but the potential for increased gain on high-resolution, short exposure time shots might be worth investigating. Daniel L. Callahan Mechanical Engineering and Materials Science Rice University, MS 321 6100 S. Main St Houston, TX 77005-1892
In addition to all of the other advice you have received, thiner sections are more difficult to stain than thicker sections are; less material to attach lead and uranium to, thus less contrast.
Geoff -- *************************************************************** Geoff McAuliffe, Ph.D. Neuroscience and Cell Biology Robert Wood Johnson Medical School 675 Hoes Lane Piscataway, NJ 08854 voice: (908)-235-4583; fax -4029 e-mail: mcauliff-at-umdnj.edu ***************************************************************
Available: SEM's, TEM's, EDS, LM's, Evaporators, Sputtercoaters, Recirculators, Ultra Microtomes, Enlargers and dark room equipment, Histology equipment, parts for some instruments and general lab equipment. SEM-TEM-Histology Services and sample preparation. e-mail your address and fax number for an Equipment List. Microscopy Labs Box 338 Red Bank, NJ 07701 fax 908 758 9142
I would like to obtain a PCVISIONplus framegrabber (PAL version). These boards made by Imaging Technologies Inc. were very popular some (6 to 8) years ago for monochrome image grabbing.
If you have one of these which you are not using and would be willing to part with it, would you please email me.
With thanks
David Vowles Electron Microscopy Unit Australian National University PO Box 475 Canberra ACT 2601 Australia Tel:(616) 2493543 Fax:(616) 2494891 Email: David.Vowles-at-anu.edu.au
Specimen blocks are very soft and are not clear but white.
I freeze yeast cells using a propane jet, freeze substitute in acetone for 4 days at -85C, replace acetone with acetone/uranyl acetate(about 0,1%) for 16 h during warm up to -35 C and wash with acetone for 2 h. Embedding: 1 h 50% HM20, 1 h 100% HM20, 16 h pure HM20, 6 h pure HM20, polymerization in gelatine cups using the Leica racks with the Leica AFS (but I use slightly smaller gelatine cups which fit directly onto Leica holder without the "Edelmann tubes"). I use relatively long times since agitation of samples is a problem in the AFS due to space limitations.
I am forwarding a message from Tony King at VG Microtech, who manufacture the Polaron range of sputter coaters, in regard to the recent question about AU/PD coatings.
Allow me to assist with your questions:
} Two questions: } 1. When a gold coated specimen is viewed in SEM at ~10k or higher, a } fine structure of gold coating becomes visible. Are there optimal } conditions of sputter coating (sputter current, time, distance } target-specimen, argon pressure, other gas than argon) to minimise the } artefact?
Without question, all of the parameters you mention are important for the high quality production of thin film for SEM observation. These are dependant on the design of the sputter head and vary from one design to the next it is impossible to give you further data without knowledge of the unit you use.
I would suggest that if you can see the coating at magnifications as low as 10k then coating is too thick! Try ten times thinner.
} 2. What is Au-Pd target? I thought that it was just an alloy one, but } a supplier of the coater says that using the alloy target is not } enough, that a coater with simultaneous sputtering Au and Pd from two } different targets is required to produce continuous coating.
The idea behind the alloy target is that the Pd acts as a barrier to the gold conglommarating into large islands which in turn obscures ultrastructure.
As there is a correlation between the work function of the metal and the sputter rate, then ideally two sputter heads with independant ontrol is the best option for Au/Pd , however, this is seen as prohibitively expensive for EM use, especially when other target amterials will give better results in some instances.
Another point of interest is that with a well designed magnetron sputter coater such as the Polaron range SC7640 system, the grain size and evenness of the coat is such that there should be no evidence of the coating with a standard SEM. This is of course dependant on the parameters you have stated earlier and a suitably thin coat.
Platinum, when used with the SC7640 will give results suitable for FEG SEM.
I hope this is of assistance to you, I have no doubt it will trigger some interesting debate on the open pages!.
Best regards Tony
Tony King Product specialist VG Microtech/ Polaron range
Tel: +44 (0)1825 746251 Fax: +44 (0)1825 768343
E&OE
Best regards, Steven Slap ******************************** Energy Beam Sciences, Inc. Adding Brilliance To Your Vision ebs-at-ebsciences.com http://www.ebsciences.com/ ********************************
Thanks to all who responded to my request for turbo pump experiences. I received over a dozen replies and there were only 1 or 2 problems=20 and these were on early models of turbo pump systems. =20 Most common remarks were; "reliable, clean, and trouble free." "Seldom need service" was also a repeated experience with the TP's. =20 Also frequently mentioned was the benefit of a vacuum system without=20 water and associated problems of corrosion, leaks etc. =20 Only negatives associated with newer turbo pumped systems was the cost= =20 of replacement or repair; many cited the wisdom of remaining on=20 service contracts.=20 =20 Possible problems with vibration and longer pump down times were=20 mentioned in a few responses but even these users with DP experience=20 preferred turbo pumped vacuum systems. =20 Thank you again for responding. =20 =20 John Humenansky Braun Intertec Corp. 6875 Washington Ave. So. Minneapolis, MN 55439 612-942-4822 =20
Thanks to all who have responded to the call for lead speciation in smelter vs natural occuring products. The information has been forwarded to the correct person and they will make there decision soon because the final report is due in 5 weeks. Thanks again.
Clarissa Vierrether The Doe Run Co. cvierret-at-misn.com
does anyone have or know of published modulation transfer functions of photographic film, especially Kodak SO-163.
Philip -- Philip Koeck Karolinska Institutet Dept. of Bioscience Novum S-14157 Huddinge Sweden Tel.: +46-8-608 91 86 Fax.: +46-8-608 92 90 Email: Philip.Koeck-at-csb.ki.se http://www_scem.csb.ki.se/pages/philip.html
MISSOURI-ILLINOIS-KANSAS MICROBEAM ANALYSIS SOCIETY & CENTRAL STATES MICROSCOPY SOCIETY
Presentations will be in the Arthur Mag Conference Center (Mag Center) of Midwest Research Institute, Kansas City, MO
APRIL 25, 1997 ====================================================== PROGRAM
8:30-9:00 Registration, Vendor Display Setup in the Mag Center.
9:00-9:10 Welcome by Dr. William P. Duncan, Director, Midwest Research Institute.
9:10-9:25 Dr. Peter Moroz, Jr., G.S. Technologies, Kansas City, MO "Microstructure of Metal"
9:30-9:45 Harold McCormick, C-K Engineering Inc., St. Louis, MO "Wearever of Metals"
9:50-10:05 Garry Crabtree, TechSpec Inc., Raytown, MO "Material Response to Deep Cryogenic Tempering"
10:10-10:30 Morning Break-Refreshments from BAR ROMA (cash cart).
10:30-10:45 Dr. Ody Maningat, Midwest Grain Products, Inc., Atchison, KS "Wheat Starch and Wheat Gluten Research"
10:50-11:05 Dr. Diane Durham, KU Medical Center, Kansas City, KS "Regeneration of Sensory Hair Cells in the Avian Cochlea-SEM Analysis"
11:10-11:25 Dr. Peter Smith, KU Medical Center, Kansas City, KS "Light and Electron Microscopic Investigation of Nervous System Plasticity"
11:30-11:45 Dr. Amit Mukherjee, KU Medical Center, Kansas City, KS "Electron Microscopic Analysis of the Polymerization of FtsZ, an Essential Cell Division Protein of E-Coli"
12:00-1:15 BUFFET LUNCH Generously provided by HITACHI, catered by Nance's Deli and Catering. Buffet includes vegetable manicotti, garden salad, garlic bread, soft drinks, etc.
1:15-1:35 Business Meetings
1:35-1:55 Paul Benson, The Nelson-Atkins Museum of Art, Kansas City, MO "Scientific Technique as Applied to Art Objects"
2:00-2:15 Marv Hart, Century Lubricants, Kansas City, KS "Lubrication and Grease Technology"
2:20-2:35 Garth Kristoff, Allied Signal, Kansas City, MO "Overview of the Technical Transfer and Solvent Substitute for Electronic Cleaning"
2:40-3:00 Afternoon Break-Refreshments from BAR ROMA (cash cart)
3:00-3:30 Michael Saba, Digital Instruments, Eden Prairie, Minnesota. "Applications in Scanning Probe Microscopy"
3:35-3:50 John Wilson, Mo. Regional Criminalistic Lab, Kansas City, MO "Capabilities of Crime Scene Investigation"
****************************************************** Complimentary lunch will be provided for all attendees by HITACHI. However, we need to know how many will be attending the luncheon, prior to the meeting. Please phone or email one of the following:
Larry Irwin 913-268-9009 Dan Kremser 314-935-5605 dkremser-at-levee.wustl.edu
} The french company ALTO MEDIA is producing a series of short movies (3 '), } based on a 'travel' into the matter.
Now it can be told... not really standard fare for this list, but I thought you all might enjoy this story.
Along the same lines, back in 1989 or so, some of the special effects for one of the Star Trek movies were done with SEM images. I think this was the first one Shatner directed, and a special-effects house called Associates & Ferran out on Long Island sold him on the use of SEM imagery because of the depth of field. If any of you remember the "planet at the end of the universe" scene, that "planet" was really a chunk of crystalline material imaged in the SEM at very low magnification.
When you hear that a film cost $50 million to make, here's how it happens. They bought a Zeiss SEM and a PGT imaging system just for the movie! A programmer who worked on camera motion-control animation systems was brought in to write stage control software to "fly" the stage through a sequence of images, which were stored on tape for post-processing. At that time, a 60MB cartridge tape was big storage. An overnight run filled up a tape and generated all of 3 seconds of animation! They did this more or less continuously for several months, and wound up using less than 10 seconds of it in the final film.
Some of the PGT engineers went to the movie the first week it opened. We got the very last credit line in the film, right behind the guys who bring the sandwiches for the actors' lunch. The people vacuuming popcorn off the theatre floor were giving funny looks to this bunch of strange folks standing in the aisles cheering at the credits 10 minutes after the movie was over and everybody else had left...
I've received lots of requests for the Microwave Staining procedure that I use for thin sections. So..... I'm sending this procedure out as a general post to everyone.
Sources for the procedure:
I use the general microwave techniques of Gary Logan, Ann Dvorak and R.T. Giberson. But, for this technique, the following is my own empirical findings. ( :-) so remember me when you reference!)
Notes on the procedure: =====================
* Section Collection:__________________ -- I collect my samples on grids, wick off excess water, and then immediately hold the back of the grid up to a 75 watt light bulb to dry. This seems to keep the sections on better. This is helpful because microwaving has a little more than the normal tendency to lift sections.
* Grid placement in stain:________________ -- When staining , I submerge the grid. Microwaving a floating grid tends to deposit uranyl acetate crystals on the grid. This is avoided by submerging the grid in the stain.
* Equipment:__________________________ --- I currently use a Ted Pella microwave, model 3440, but I've used a standard kitchen microwave before with good results.
--- Porcelain shallow well evaporating dish ( 12 wells)
--- Syringes &Filters for the stains used.
*Stains:_______________________________ --- Saturated (10%) Uranyl Acetate ( aq), less than 10 days old. Make in very small quantities. ( Our water , building or something is odd, this is only how long our aqueous solution lasts)
--- Venable's lead stain (John Venable, Richard Coggeshall,"A Simplified Lead Citrate Stain."The Journal of Cell Biology, 1965, Vol 25, pp407-408)
========================================= Procedure: ** Microwaving is actually only in the Uranyl acetate step.
1. For no more than 8 grids at a time, place about .25ml of filtered U.A. in a well for each pair ( identical pair) of grids, for a max of 4 wells that have U.A.
2. Fill all other wells with .25-.5 ml of water.
3. Place the grids , section side down, into and to the bottom of the drop. Be sure grids do not overlap.
4. Prewarm microwave that has 2 --300 ml water baths.
( Each Microwave oven is different, may need to test where best placement is. I use one water bath at 9 o'clock and one at 12 o'clock. Test using fresh cool water in the 2 beakers, place a liquid crystal sheet {Ted Pella} on the bottom, use a temp range of 35-40 or 40-45C......... look for the non-heated spots , this is where to place your sample)
5. Replace water in the water baths.
6. Place the evaporation plate in the "cool" spot of the microwave.
7. Microwave for 33 seconds and let plate set there for 6-8 minutes. Change water baths.
8. Repeat step 7 for a total of 4 times The total incubation time should be at least 20 min, but less than 30.
9. Leave the grids in the stain as you wash them one by one in warm water.
10. Proceed with normal lead staining. ( I use the Venable's for 40 seconds - Room temp).
FAQ's: ~~~ WHY MICROWAVE IF IT MAY TAKE 20 MINUTES ANYWAY?
The improvement in staining is really obvious. Also, I use a lower contrasting resin, and so this helps a lot. For things you are having trouble staining, this helps a lot.
~~~ CAN ONE OVER STAIN???~~~~~~~~~~~~~~~~~~~~~~~~~~~
Yes! Do a pilot study first. If you are working with a resin mixture that stains well, you may find that only 6-7 minutes is more than enough. I had a mixture at one time that did very well at 5 minutes only in total incubation time. I use what I do now for the way the mixture cuts.
~~~ WHY NOT JUST USE ONE BIG LONG MICROWAVE SESSION?
--The stain gets too hot, it will evaporate, and causes precipitation in the wells and on the grid. -- The sections will tend to come off if times of 1-2 min of microwaving are used. This is actually the method I started out with, and it doesn't work as well.
~~~WHAT ARE THE MOST COMMON PROBLEMS TO WATCH OUT FOR?
-- Overheating , will dry out the stain, and put stain ppt on the grid. -- Lifting sections, some batches of epoxy stick better than others, helps to dry the section on the grid with a light bulb, and to gently lower the grid in section side down. -- Over staining......was a new possibility for me! -- Lou Ann Miller Center for Microscopy & Imaging College of Veterinary Medicine Dept. of Veterinary Biosciences University of Illinois Rm 1108 Basic Sciences Bld 2001 S Lincoln Ave. Urbana, Illinois 61802
Phone: 217-244-1566 email: lamiller-at-uiuc.edu
Center for Microscopy & Imaging Home page: http://www.cvm.uiuc.edu/MicImagLab/MicImagLab.html
Central States Microscopy Society: http://www.cvm.uiuc.edu/Homepages/LouAnnMiller/CSMS/csms.html
Personal Home Pages: http://www.cvm.uiuc.edu/Homepages/LouAnnMiller/LAM.html
MISSOURI-ILLINOIS-KANSAS MICROBEAM ANALYSIS SOCIETY & CENTRAL STATES MICROSCOPY SOCIETY
Presentations will be in the Arthur Mag Conference Center (Mag Center) of Midwest Research Institute, Kansas City, MO
APRIL 25, 1997 ====================================================== PROGRAM
8:30-9:00 Registration, Vendor Display Setup in the Mag Center.
9:00-9:10 Welcome by Dr. William P. Duncan, Director, Midwest Research Institute.
9:10-9:25 Dr. Peter Moroz, Jr., G.S. Technologies, Kansas City, MO "Microstructure of Metal"
9:30-9:45 Harold McCormick, C-K Engineering Inc., St. Louis, MO "Wearever of Metals"
9:50-10:05 Garry Crabtree, TechSpec Inc., Raytown, MO "Material Response to Deep Cryogenic Tempering"
10:10-10:30 Morning Break-Refreshments from BAR ROMA (cash cart).
10:30-10:45 Dr. Ody Maningat, Midwest Grain Products, Inc., Atchison, KS "Wheat Starch and Wheat Gluten Research"
10:50-11:05 Dr. Diane Durham, KU Medical Center, Kansas City, KS "Regeneration of Sensory Hair Cells in the Avian Cochlea-SEM Analysis"
11:10-11:25 Dr. Peter Smith, KU Medical Center, Kansas City, KS "Light and Electron Microscopic Investigation of Nervous System Plasticity"
11:30-11:45 Dr. Amit Mukherjee, KU Medical Center, Kansas City, KS "Electron Microscopic Analysis of the Polymerization of FtsZ, an Essential Cell Division Protein of E-Coli"
12:00-1:15 BUFFET LUNCH Generously provided by HITACHI, catered by Nance's Deli and Catering. Buffet includes vegetable manicotti, garden salad, garlic bread, soft drinks, etc.
1:15-1:35 Business Meetings
1:35-1:55 Paul Benson, The Nelson-Atkins Museum of Art, Kansas City, MO "Scientific Technique as Applied to Art Objects"
2:00-2:15 Marv Hart, Century Lubricants, Kansas City, KS "Lubrication and Grease Technology"
2:20-2:35 Garth Kristoff, Allied Signal, Kansas City, MO "Overview of the Technical Transfer and Solvent Substitute for Electronic Cleaning"
2:40-3:00 Afternoon Break-Refreshments from BAR ROMA (cash cart)
3:00-3:30 Michael Saba, Digital Instruments, Eden Prairie, Minnesota. "Applications in Scanning Probe Microscopy"
3:35-3:50 John Wilson, Mo. Regional Criminalistic Lab, Kansas City, MO "Capabilities of Crime Scene Investigation"
****************************************************** Complimentary lunch will be provided for all attendees by HITACHI. However, we need to know how many will be attending the luncheon, prior to the meeting. Please phone or email one of the following:
Larry Irwin 913-268-9009 Dan Kremser 314-935-5605 dkremser-at-levee.wustl.edu
I'm looking for carbon EM calibration grid from SPI model 411CG-AB but couldn't find any information about SPI.Does anyone knows they phone # or www address? Thanks.
**************************** * Maoxu Qian, Ph.D. * * Dept of MSE, box 352120 * * University of Washington * * mxq-at-u.washington.edu * * (206)543-1514(phone) * * (206)543-3100(fax) * ****************************
SUMMER I 1997 COURSE ANNOUNCEMENT - Transmission Electron Microscopy (BIO. 221-Section BA)
NASSAU COMMUNITY COLLEGE, Garden City, Long Island, New York
A five week, Summer Session I 1997 semester, course in Biological Transmission Electron Microscopy is being offered by the Biology Department of Nassau Community College. This is a 4 credit course offered four days per week (Monday through Thursday) between the hours of 8:00 am and NOON. Classes will begin on May 27 and end on June 26, 1997.
This is a "hands-on" course emphasizing biological specimen preparation, ultra-thin sectioning involving block trimming, glass knifemaking and operation of the ultramicrotomes (Sorvall MT-2B and LKB Ultrotome III), thick and ultra-thin section mounting and contrast staining (UA and Pb citrate), grid support films (formvar, carbon), student operation of the TEM (Hitachi HS-8, Philips EM 300) and production of electron micrographs through the process of black & white photography, and electron micrograph analysis. Students will work on a chosen sample(s) with the goal of producing a portfolio of high quality TEM photomicrographs of that sample(s).
The course is widely transferrable and the cost per credit is reasonable at $78 per credit.
More information about the Bio-Imaging Center at NCC, course descriptions and syllabi, and the humble beginnings of a student gallery of EM photomicrographs is available at our web site. The URL is {http://www.sunynassau.edu/webpages/biology/becks.htm} .
Interested individuals should register as soon as possible since the course is limited to a total enrollment of ten (10) students.
If you have further questions, you should e-mail me directly at the address below.
Stephen J. Beck Associate Professor Bio-Imaging Center/Electron Microscopy Department of Biology Nassau Community College Garden City, NY 11530 Voice Mail: (516) 572-7829 Email: {becks-at-sunynassau.edu} URL: {http://www.sunynassau.edu/webpages/biology/becks.htm}
} does anyone have or know of published modulation transfer functions } of photographic film, especially Kodak SO-163.
There has not been much published lately... I believe that the most recent data comes from K. H. Downing and D. A. Grano, "Analysis of photographic emulsions for electron microscopy of two dimensional crystalline specimens," Ultramicroscopy, 7, 381-404 (1982).
-- Best Regards, John Minter
Eastman Kodak Company Phone: (716) 722-3407 Analytical Technology Division FAX: (716) 477-3029 Room 2112 Bldg 49 Kodak Park Site email: minter-at-kodak.com Rochester, NY 14562-3712 calendar: via PROFS
To all those who asked for a copy of a book chapter discussing staining: I am leaving for NYC for a week shortly. I will honor your request upon my return, April 22. Bye, Hildy
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Postdoc Position starting Oct. 1, 1997 at ANL-West
Argonne National Lab - West has an opening for a postdoc position to investigate irradiation damage in stainless steels from the EBR-II reactor. The project will mainly involve studying the defect structure and grain boundary segregation in up to ten different reactor core components. STEM will be the main technique used; however SEM, micro-hardness testing, and mechanical testing will be required occasionally.
Equipment available: JEOL 2010 STEM (LaB6) Zeiss 960A SEM Wide range of sample preparation equipment all capable of preparing radioactive samples.
Microscopy Skills Desired: Dark Field Imaging, EDS, Electron Diffraction, STEM, CBED
General Skills Desired: Experience handling/preparing radioactive samples Excellent verbal and written communication
Education Required: Ph.D. in Materials Science or Nuclear Eng. specializing in irradiation effects
This position will start Oct. 1, 1997.
Send Resumes to Brad Storey P.O. Box 2528 Idaho Falls, ID 83403
I would like to know what sort of probe current I should expect to measure at the specimen in a JEOL 2000fx TEM using a LaB6 filament, 200kV, 120 micron condenser aperture and spot sizes 1L to 6L. Under these conditions I measured the currents (using the faraday cup in our Gatan analytical specimen holder) to be:
1L 170nA 2L 66nA 3L 40nA 4L 7nA 5L 1nA 6L 0.3nA
Are these values reasonable? Cheers,
Mark Blackford TEM Group Materials Division, ANSTO PMB 1, Menai, N.S.W. Australia 2234 Phone 61 2 9717 3027 Fax 61 2 9543 7179
Disclaimer: The views expressed in this E-mail message do not necessarily represent the official views of ANSTO from which this message was conveyed.
can anyone tell me what purity of SF6 is required for use in TEM HT tanks and guns. I have a JEOL 2000 fx which was converted to SF6 in late 1994. We operate this machine at 200kV almost exclusively. At the time of the modification there was only one grade of SF6 available in Australia, as far as I could determine. I believe this was 99.8% pure. It has recently come to my attention that this may not be good enough. Before I go to the trouble and expense of sourcing higher purity SF6 I would like a definitive answer to the the question "how pure is pure enough?". JEOL Australia hasn't been able to help.
I look forward to any insight you can provide. Please respond to the listserver as there are several others in Australia that are keenly interested in this as well. Thanks,
Mark Blackford TEM Group Materials Division, ANSTO PMB 1, Menai, N.S.W. Australia 2234 Phone 61 2 9717 3027 Fax 61 2 9543 7179
Disclaimer: The views expressed in this E-mail message do not necessarily represent the official views of ANSTO from which this message was conveyed.
We have a JEOL 2010 and we use 99.995% purity SF6. I order it from a company which calls it VLSI grade , but I seem to recall that they used to call it Instrument grade.
We order a C-size cylinder (10 lb). Last time I ordered was about 2 years ago and we still have the cylinder with a fair amount of gas in it.
My background is in EM of biological, medical and food samples. I now have the opportunity to expand my skills and work on a set of materials samples.
My question is:
What are the standard methods for assessing surface degradation of materials samples (especially plastic polymers)? Are there macroscopic as well as microscopic methods which can give statistically "good" descriptions of the extend and type of degradation of such surfaces?
Thanks for any help (methods, references, review papers, etc.) you can provide. Please contact me offline.
Paula.
Paula Allan-Wojtas Food Microstructure Specialist Agriculture and Agri-Food Canada Atlantic Food and Horticulture Research Centre Kentville, Nova Scotia,Canada B4N 1J5
} For Sale: JEOL JSM T-200 scanning electron microscope. Purchased in } 1984; used about 50 hours, total time. Includes chiller and a small } sputter coater. $1500 OBO. Contact Dr. Jon Martin at 912/752-4060. } Located at Mercer University School of Medicine, Macon Georgia.
E-mail contact: HORST_MN-at-Mercer.EDU ******************************************************* G.W. Erdos, Ph.D. Phone: 352-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax: 352-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 http://www.biotech.ufl.edu/~emcl/ Home of the #1 Gators ***** "Many shall run to and fro, and knowledge shall be increased" Daniel 12:4
From the first superconducting magnet to the next generation of semiconductor chips, Oxford Instruments have always had an eye to the future - a future of scientific, human and commercial progress through the development of technology and people.
The Microanalysis Group is a highly successful international business within Oxford Instruments plc. We develop and manufacture high quality instrumentation for X-ray microanalysis and imaging of materials in electron microscopes. We are a world market and technology leader certified to BS EN ISO9001.
Continued expansion has led to an immediate vacancy in the Software R&D department for a researcher to improve our understanding of the physics of excitation & detection and develop innovative software algorithms to extend the range of microanalysis applications.
The successful candidate will have: - A relevant science degree or PhD with a good background in numerical methods and basic statistical theory. - Experience of EDX and/or WDX microanalysis and SEM theory and operation. - Programming skills in C and possibly Visual Basic on a PC platform.
In return for your commitment, we offer the full benefits package you would expect form a large plc. Career prospects are excellent and training will be tailored to the needs of the successful candidate.
If you are interested, please send a full covering letter of application and a comprehensive CV, quoting reference MRD01 on the envelope to:
Helen Bacon Personnel Manager Oxford Instruments Microanalysis Group Halifax Road High Wycombe, Bucks. HP12 3SE ENGLAND
or e-mail: helen.bacon-at-oxinst.co.uk -- Oxford Instruments MAG Software R&D
The Advanced International Immunofluorescence Course is a post-doctorate theoretical/practical course, with propedeutical lectures and practical stages on traditional and confocal immunofluorescence microscopy and image and ion analysis. The course will take place in Gargnano (Lake of Garda) from 7 to 10 October 1997. Further information and registration details will be found at the following Web address
http://imiucca.csi.unimi.it/endomi/ACIF.html
Thank you Paolo Castano _______________________________________________________________________
Prof. Paolo Castano UNIVERSITY OF MILAN INSTITUTE OF HUMAN ANATOMY - CHAIR OF HUMAN ANATOMY FOR PHARMACY Via Mangiagalli, 31 - 20133 Milan (Italy)
Dear fellow microscopists, Would anyone be interested in helping with a project on TEM at a high school level? My daughter Kelly, is doing research paper for intergrated sciences and is asked to find help from experts in the field (Mom's don't count) who can mentor her regarding some of the basic principles involved in TEM. She must cover chemistry, physics, mechanics and biology in relation to her topic. Her paper (due mid May, but preparing now) will cover some of these questions:
Why electrons are a better source than light for resolution? How does wavelength effect resolution? What makes Tungsten a good electron source, and how are the electrons generated? How does a a magnetic lens work? Why is a vacuum needed for operation? If electrons are invisible, how is the image generated off of the screen? How are images made on the film and then chemically developed? How does the eye focus and transmit this information to the brain?
If anyone would like to take a crack at any of the above with an explaination aimed toward the H. S. level (16yrs old. but fairly advanced science knowledge), it would be most appreciated. Of course, your mentoring would be acknowledged in her references. Please send all e-mail regarding this off line to me and I will forward if to Kelly. Thanks for promoting EM to young scientists! Linda Fox lfox1-at-wpo.it.luc.edu
It does seem strange that JEOL doesn't know what purity SF6 it prefers for its tanks. One of the other microscope vendors specified 99.9% or better in an installation guide for one of our TEM's.
} can anyone tell me what purity of SF6 is required for use in TEM HT tanks } and guns. I have a JEOL 2000 fx which was converted to SF6 in late 1994. } JEOL Australia hasn't been able to help.
Russell E. Cook Scientific Associate Electron Microscopy Center for Materials Research Argonne National Laboratory Building 212 9700 South Cass Avenue Argonne, IL 60439 (630)252-7194 FAX: (630)252-4798
The New England Society for Microscopy (NESM) announces its 14th ANNUAL SPRING SYMPOSIUM to be held at the Marine Biological Laboratories in Woods Hole, Massachusetts on May 9 & 10, 1997. Cohosted by the Connecticut Microscopy Society (CMS), the Metropolitan Microscopy Society (MMS) and the New York Society for Experimental Microscopists (NYSEM).
PROGRAM Friday, May 9th
12:00 pm Registration: Swope Center
1:00 pm Welcome: Lillie Auditorium
Session I Chairperson: Philip Leopold, NYSEM
1:10 pm "System Integration in Light Microscopy" Kenneth Spring, National Heart, Lung and Blood Institute, Bethesda, Maryland
1:50 pm "Diagnosis of Strange and Exotic Animal Diseases" Douglas Gregg, Foreign Animal Disease Diagnostic Lab, NVSL,USDA
2:30 pm "EM Site Magnetic Field Surveys and Solutions" Curt Dunnam, Cornell University, Linear Research Associates
3:10 pm Afternoon Break
Session II Chairperson: Joe Antol, CMS
3:30 pm A brief talk on "Amine Catalysts and Embedding Media" Jose Mascorro, Tulane University, MSA/LAS Director
3:50 pm "mRNA Localization and Cellular Morphogenesis" Gary Bassell, Dept. of Anatomy, Albert Einstein Medical School
4:30 pm "Quantitative Determination of Elemental Segregation at Interfaces in Solids" Tony Garratt-Reed, Center for Materials Science and Engineering, M.I.T.
5:30 pm Cocktails and Dinner: Swope Center
7:30 pm "Tracking the Giant Bluefin Tuna in New England Waters" Molly Lutcavage, New England Aquarium Edgerton Research Laboratory
Saturday, May 10th 7 to 8:00 am Breakfast: Swope Center
Session III Chairperson: Philip Flaitz, MMS
8:30 am "Ion Beam Milling Materials with Applications to TEM Specimen Preparation" Ron Anderson, IBM Analytical Services Group
9:10 am "Interaction of Viable Listeria Monocytogenes with Host Cell MHC Class II Compartments and Low pH Compartments" Paul Webster, Center for Cell Imaging and Department of Cell Biology, Yale School of Medicine
10:00 am Commercial Exhibits and Posters: Swope Center
12:30 pm Presentation of Poster and Photos-As-Art Awards, Door Prizes and "Nobska Light" Art Raffle Drawing: Poster Area, Swope Center
1:00 pm Lunch with Short Tour of MBL: Swope Center
2:00 pm 90-minute Discovery Cruise aboard the R/V Patriot II Tickets must be reserved in advance
This program is supported in part by a grant-in-aid from the Microscopy Society of America.
TO REGISTER, contact L. Kirstein at 104365.3522-at-compuserve.com. Advance registration is required. Deadline for cocktail/dinner reservations is April 25, 1997.
Evex Analytical is offering an X-ray detector enginer in its Princeton, New Jersey office.
Please foward resume to
Human Rresources Evex Analytical 857 State Road Princeton, NJ 08540
Candidates should have: - Experience of EDX and/or WDX microanalysis - A good background in numerical methods and basic statistical theory. - Programming skills in C, Visual Basic, Visual C++, Java, Active X
Please foward resume to
Evex Analytical 857 State Road Princceton, NJ 08540
The easiest, and most reliable method is to } buy commercially titrated NaOH (1 N ). No worries then. No need to try } a pH meter. Perfectly reliable.
This thread has offered some thought provoking ideas on the preparation of Reynold's. I have one question: how stable is commercially titrated NaOH? Doesn't a solution of NaOH absorb CO2 from the atmosphere and form sodium carbonate? Wouldn't this lead to a change in molarity with time and promote the formation of lead carbonate ppt on the sections?
Thomas E. Phillips, Ph.D. Associate Professor of Biological Sciences Director, Molecular Cytology Core Facility 3 Tucker Hall University of Missouri Columbia, MO 65211 (573)-882-4712 (voice) (573)-882-0123 (fax)
} can anyone tell me what purity of SF6 is required for use in TEM HT tanks } and guns.
We have a JEOL 4000, which uses Commercial Grade SF6, and an HVEM, in which we have traditionally used Instrument Grade SF6. Due to the re- cent price explosion for SF6, we have been investigating whether we can use the Commercial Grade SF6 in the HVEM as well. The major difference between the two grades is that there is more N2 and O2 in the Commercial Grade. The HVEM has a gas storage facility with several filters and a molecular seive through which SF6 can be cycled. I think that much of the N2 and O2 can be eliminated by venting a small amount of SF6 to the air before putting the rest in the tank. You'd have to calculate the dielectric constant and know the relevant spark gap widths, etc. to be sure if Commercial grade would work for you, but our conclusion at this point is that it works for the 400 kV instrument, and should be OK for the HVEM as well. (We are still going to analyse both grades for im- purities.) Good luck. Yours, Bill Tivol
} What are the standard methods for assessing surface degradation of } materials samples (especially plastic polymers)? Are there macroscopic } as well as microscopic methods which can give statistically "good" } descriptions of the extend and type of degradation of such surfaces? } Dear Paula, I recently ran across a good review "Electron Microscopy in Polymer Science" by G.H. Michler, Applied Spectroscopy Reviews, vol 28(4), pp 327-384 (1993). This paper discusses surface characteriza- tion and many other topics. Additionally, I suggest STM or AFM as possibilities, but I don't have any referrences for them. Perhaps comparing the specular vs diffuse reflectivities of polymer surfaces before and after damage would be informative for degradation features ~ 1 micrometer in size. Good luck. Yours, Bill Tivol
Just a quick note to let you all know that that Microscopy Listserver Archives are now on-line. I have made accessible all Email postings covering the period Oct.1993 through Mar.1997 and will update the archive monthly.
The index is not directly searchable, however, it is chronologically sorted by Month and Year.
If you download a given month's postings then you can search the downloaded WWW page for any keyword/phrase that you wish by using the native search/find option of your WWW Browser. (In NetScape this is located in the Edit Pull Down Menu and is called FIND).
You may access the archive at the MSA WWW site.
http://www.msa.microscopy.com
just follow the links to the Reference/Educational Activities Page.
I'll get around to putting together a completely searchable index sometime in the forseeable future, but for now this will go a reasonable way to letting everyone find "old messages and postings" and all the other miscellaneous requests I receive for information.
Way back in the mid 1980's there was a book called "Thin Is In" and it dealt with staining of samples embedded in GMA. Are there still copies out there? Or can someone direct me to where I can get ahold of the book so I can phototcopy pertinent pages? I'm sorry, but I don't remember who the author was.
Thanks in advance for all you help.
Paula = )
Paula Sicurello UC Berkeley Electron Microscope Lab psic-at-uclink4.berkeley.edu
A while ago I remember reading that not only should the lead stain be made up at the proper pH (12), but both the lead citrate powder (for Venable's) and the NaOH need to be fresh. I bought new reagents (esp. C02-free NaOH solution) and got much cleaner, stronger staining.
Also, I never had very great uranyl acetate staining. When I was doing work on collagen, I switched to a tannic acid/uranyl acetate stain (ref: Kajikawa et al., 1975, J. Electron Microsc. 24:287-289) and I have since used it on most everything. Again, it seems to give stronger, cleaner staining than the UA alone. This stain (a modification of the one in the reference) must be made fresh on the day of use, so I usually make fairly small volumes. I pre-weigh 0.04 gm tannic acid into a few tubes. On day of use add 6.8 ml distilled water to one tube, mix and hold in hands or place in oven about 5 minutes to warm. Mix again, then add 200 ul of 2 % aqueous uranyl acetate. Mix and spin down or put through syringe filter before use. Stain 10-15 minutes and rinse in water before going to the lead stain.
Another reason I like the above solution is that, according to my calculations anyway, if I start with depleted uranyl acetate to make the 2%, the final tannic acid/UA stain solution does not contain enough radioactivity to be considered radioactive. So I don't have to worry about radioactive staining dishes, forceps, etc. etc. as long as I am careful with the original UA and 2% solutions. Of course most of the materials I use are disposable, and the waste profiles are designated to contain some UA so it really doesn't matter alot. But it makes me feel better to limit usage of the stuff.
Now if I could just learn how to stain sections on formvar-coated grids without getting lots of folds and dense pockets!
Good luck,
Karen Zaruba
} This should be a simple procedure but I am having a terrible time trying to } stain some Embed 812 embedded tissues with uranyl acetate/lead citrate. The } tissue is from frog tadpoles, fixed in glutaraldehye/paraformaldehyde and } postfixed in osmium. I have tried staining for 5-20 minutes with saturated } solution of UA in ethanol, followed by 1-5 minutes in lead citrate } (Venable-Coggeshall formulation). The sections look no different from } unstained specimens. } } I'd appreciate any suggestions about what I might be doing wrong. } } } Gary Radice, Associate Professor gradice-at-richmond.edu } Department of Biology 804-289-8107 (voice) } University of Richmond VA 23173 804-289-8233 (FAX) }
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Life Sciences Sector Lab Reply: kszaruba-at-mmm.com 3M Company 3M Center 270-1S-01 Phone: 612-737-2971 St. Paul, MN 55144-1000 Fax: 736-1519
These opinions are my own and may not represent those of 3M.
I have recently been asked to take over an ICP. Since this newsgroup has been so helpful with microscopy issues I was wondering if anyone knows of a similar newsgroup dedicated to ICP spectroscopy.
Could someone please give me the name of a book which would have TEM pictures of paper which has been prepared with ultramicrotomy. Or if they have one or two thay would be willing to send to me privately I would really appreciate it.
For our JEOL 2010, Jeol specify a purity of 99.99% or better in their maintenance manual. We however have 99.995% purity obtained from Scott Specialty Gases, Fremont.
Best Regards,
Keith. ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Dr. K. Moulding.
Materials Characterisation and Preparation Facility Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong.
Below is a request for instructions and/or parts for a cryostat from some colleagues.
Thanks in advance for your help!
Tina **************************************************************************** * Tina (Weatherby) Carvalho * tina-at-pbrc.hawaii.edu * * Biological Electron Microscope Facility * (808) 956-6251 * * University of Hawaii at Manoa * http://www.pbrc.hawaii.edu/bemf* **************************************************************************** Message:
We have recently obtained a surplus cryostat, a Milles Scientific Microtome Model 4553. The unit chills well and the microtome is mechanically in good shape. Unfortunately, it does not have instructions, an antirolling plate or specimen stubs. Does anyone know a source for these items? We have been unable to locate a phone number or address for Milles Scientific. Perhaps they have gone out of business or merged with another company. Perhaps someone has surplus parts for this model we could acquire. Your assistance would be appreciated.
In nearly 30 years of preparing lead citrate according to Reynolds I have experienced no problems in using a stock sodium hydroxide solution (1M) which was made up from pellets. However, this solution must be freshly made up. Is it because I always use Reynolds in its concentrated form that I do not experience any problems?
Rob
Robin H Cross Director : EM Unit, Rhodes University, Grahamstown, South Africa eurc-at-giraffe.ru.ac.za - tel: +27 461 318168 - fax: +27 461 24377
Materials Analysis Group, Philips Semiconductors, has an immediate opening for an experienced XRD analyst.
Responsibilities include operating XRD and scanning acoustic microscopy on bulk/thin film specimens and packaged devices, respectively, primarily for external customers on a commercial basis. Analytical support work for internal customers on silicon IC production issues will also be required.
Qualified candidates must possess a Ph.D. or M.S. degree in a field such as Materials Science or have equivalent expertise. They must have extensive hands-on experience in operating XRD equipment and in providing analyses to customers, preferably as a member of an analytical services laboratory. Familiarity with IC processing would also be advantageous.
Equipment available includes a Siemens D500 with grazing incidence and thin film reflectivity attachments, and a Sonoscan C-SAM. Acquisition of a thin film diffractometer is under consideration. The laboratory is also well equipped for SIMS, Auger, ESCA, TEM, SEM AFM, FIB, Raman, FTIR, etc.
Located in the heart of Silicon Valley, 40 miles south of San Francisco, Philips Semiconductors offers a generous benefits package that includes life, health and dental insurance, a pension plan, and a company-matched savings and investment plan.
For confidential consideration, send your resume to: Alan Morgan, Materials Analysis Group, Philips Semiconductors MS 65, 811 E. Arques Avenue, Sunnyvale, CA 94088: FAX (408)991-4801.
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Paula, Thin Is In: Plastic Embedding of Tissue for Light Microscopy. Burns, William A. & Bretschneider Ann. 1981. Educational Products Division. American Society of Clinical Pathologists. Chicago. ISBN 0-89189-083-1
I have a copy, so send your fax number, what you would like copied I'll get them to you. Ian.
Fellow microscopists I am doing some TEM of cultured lymphocytes. All membranes seem to be absent. There are halos where membranes should be but no membranes. So far I have tried 2.5 glut in 0.1M cacodylate 2.5 glut in 0.2M cacodylate (caused shrinkage) 4 paraformaldehyde 1 glut in 0.1M cacodylate.
Post fixing in Osmium dehydrating in ethanol and embedding in Spurr
I am going to try making up the fix in the culture medium next.
Has anyone any thoughts of anything else to try. I would be particularly interested in the use of Ca+ Mg+ and sucrose.
Many thanks
Chris Chris Gilpin Biological Sciences Electron Microscope Unit G452 Stopford Building Oxford Road Manchester M13 9PT phone +44 161 275 5170 fax +44 161 275 5171
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America Like Rob I've been making Reynold's lead citrate since I was a boy. I use NaOH pellets but degas the distilled water by sonicating it for a few minutes before making the stain.
A discussion of SF6 took place on this list back in December. At that time I received an off-line message from John Giles about Dilo Company. Dilo sells (and rents) a system that reclaims the SF6 in your tank, purifies it and pumps it into storage tanks for reuse. Dilo's business is primarily with big consumers of SF6 - power companies, but they are knowledgeable about SF6 purity and applications. What interested me was they sell a small system for ~$5,000, which is what a tank of SF6 costs in some places (like the U.P. of Michigan).
I have not used their system. Has anyone else?
You can reach them at;
Dilo Company, Inc. 231A Douglas Rd.., Unit 5 Oldsmar, FL 34677 813-855-1448 http://www.dilo.com dilo-at-cent.com
I have no interest in Dilo Company other than as a consumer. Owen P. Mills Michigan Technological University Metallurgical & Materials Engineering Rm 512 MME Building Houghton, MI 49931 906-487-2002 906-487-2934 FAX opmills-at-mtu.edu
At 03:49 PM 4/15/97 -0700, Jill Craig asked: } I have recently been asked to take over an ICP. Since this newsgroup } has been so helpful with microscopy issues I was wondering if anyone } knows of a similar newsgroup dedicated to ICP spectroscopy. } I have used Netscape search engines with no luck.
I don't know the specific answer, but the best way to find lists such as this is to use http://www.liszt.com
Best regards, Steven Slap ******************************** Energy Beam Sciences, Inc. Adding Brilliance To Your Vision ebs-at-ebsciences.com http://www.ebsciences.com/ ********************************
Is anyone out there looking for a used Duo Mill for materials TEM sample prep? Excellent working condition and a very reasonable price. If interested contact Kim Jones 303-421-3182 or send e-mail kjonesVMS-at-aol.com
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Chris, For cultured cells I routinely fix in 2% glut/0.1M. cacod and post fix with 1% OsO4/0.1M. cacod, embed in Araldite, then stain sections with Ua/Pb with no problems. What are your fixation conditions - monolayer, pellet of cells. From your description sounds like the OsO4/Pb side of things is at fault. I wouldn't add fixatives to the culture medium, depending on its constituents you might have a Canazarro type reaction and fix the medium as well as the cells. Ian.
Me, too. I boil my water, cool it a little, dissolve the pellets and make the whole mess warm. I use the fancy carbonate-free pellets. I don't keep the NaOH, either-just add it to my buffer-adjusting stock. I've never tried sonicating but sounds good and easy. I wouldn't dream of sticking my pH electrode in my lead-I use a fresh piece of close-reading paper. Seems to work just fine. Grace
Been using 99.8 SF6 for several years in our JEOL 2010. Generally works well. We have groups working at 100 & 200KV. When not used at 200kv for several mo., some dark current instability is exibited when returning to 200KV. With a bit of patients we are back in business. The stability problem ceases to exist with regular use at 200KV.
I've noticed a lot of people seem to be using Reynold's Pb stain. In the past (20 years ago) I used Reynold's also. There seemed to be too many problems with the staining. I started using lead citrate to make up my stain. Consistantly good results. Here's how if anybody is interested.
Distilled water in clear glass storage bottle...........about 80 ml lead citrate.................................................0.4gm
Stir vigorously on magnetic stirrer for several minutes avoiding bubbles.
10N NaOH (I buy from Fisher already made up. 100 ml bottle).....0.2ml
Continue to stir until solution is clear. Bring up to 100ml. Store in 4C refrigrator. Good for up to 6 months. Avoid shaking it.
Stain grids for 4 minutes, rinse grids.
As far as flat beer goes, try adding CO2 under vacuum and let it sit for a day.
Peace through Christ,
Phil Rutledge, Director e-mail: prutle1-at-gl.umbc.edu Center for Microscopy voice: (410) 455-3582 UMBC Dept. of Biology fax: (410) 455-3875 Catonsville, MD 21228 ///// / / / / /////// /////// / / /////// /////// / / / / / / / / / / / / /////
Would freshly distilled water be equivalent to degassing via sonication ?
Leo
On Wed, 16 Apr 1997, Ian Montgomery wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } ------------------------------------------------------------------------ } } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } } -----------------------------------------------------------------------. } } } } Good morning Reynolds users } } } } In nearly 30 years of preparing lead citrate according to } } Reynolds I have experienced no problems in using a stock } } sodium hydroxide solution (1M) which was made up from pellets. } } However, this solution must be freshly made up. Is it because I } } always use Reynolds in its concentrated form that I do not } } experience any problems? } } } } Rob } } } Like Rob I've been making Reynold's lead citrate since I was a boy. } I use NaOH pellets but degas the distilled water by sonicating it for a few } minutes before making the stain. } } Ian. } } }
To All I have been doing TEM since 1970, and have always used NaOH pellets (CP grade - which do contain about 0.5 - 1% Na2CO3). I make up a 10N solution in small aliquots (~20 ml), and dump it when I see crystals of sodium silicate (NaOH dissolves glass). I use this 10N solution to add to the powdered Pb citrate in dH2O. I make up Pb citrate in 100 ml quantities and dump it when it turns cloudy. Usually it keeps for months, unless someone forgets to tighten the cap, which is not uncommon in a central service lab. What I have learned in nearly 30 years of TEM, and many years in other microscopy techniques is that there are very few empirical rules. I have been in very fussy labs that used aged Pb citrate in bottles that were so coated with precipitate that they looked out of a sunken pirate ship, and have been in other labs that make up Pb citrate fresh, and filter it. Whatever works. Bruce Cutler, Microscopy Lab, Univ. Kansas, Lawrence KS
I am looking for information and possibly a handbook on an American optical Micro-Star Illuminator model 1872 (I am new to this scope). This microscope was discontinued and replaced by another microscope - the EpiStar metallurgical microscope which (I believe) has been discontinued itself.
I would like to hear from anyone who has used this scope. I am also in need of objectives (the optical kind).
Chris: What are the times and temperatures? Tissue cultured cells require only about 5 minutes in each fixative at 20 degrees C or at the most for 15 minutes in ice, unless you use much lower concentrations. Overfixing or storage in con. ethanol removes lipids and hence membranes. Badly Os overfixed tissues show dark cytoplasms surrounded by "white" cell membranes. Regards Jim Darley
ProSciTech Microscopy PLUS PO Box 111, Thuringowa QLD 4817 Australia Phone +61 77 740 370 Fax: +61 77 892 313 Great microscopy catalogue, 300+ Links, MSDS ************************ http://www.proscitech.com.au
} I am doing some TEM of cultured lymphocytes. All membranes seem to } be absent. There are halos where membranes should be but no } membranes. } So far I have tried } 2.5 glut in 0.1M cacodylate } 2.5 glut in 0.2M cacodylate (caused shrinkage) } 4 paraformaldehyde 1 glut in 0.1M cacodylate. } } Post fixing in Osmium dehydrating in ethanol and embedding in } Spurr } } } I am going to try making up the fix in the culture medium next. } } Has anyone any thoughts of anything else to try. I would be } particularly interested in the use of Ca+ Mg+ and sucrose. } } } Many thanks } } Chris } Chris Gilpin } Biological Sciences Electron Microscope Unit } G452 Stopford Building } Oxford Road } Manchester } M13 9PT } phone +44 161 275 5170 } fax +44 161 275 5171
I and other collegues have been trying fruitlessly to e-mail Kevex and when I tried to find their web page it was not there either. Is this just a faulty computer or have they been restructured out of existence? Does anyone out there know?
No, freshly distilled water is not necessarily degassed. Air can redissolve nicely as the water drips down from the condenser into the collecting flask.
But as an alternative to sonification, you can try just plain BOILING. This reduces the amount of dissolved gas significantly. Then just let it sit. Don't shake it with air, try not to pour it too much. This worked for us with a different degassing problem a couple years ago.
Cindy Bennett Hoechst Diafoil GmbH Wiesbaden, Germany bennett-at-msmhdg.hoechst.com ---------- Von: Leo Marin An: Ian Montgomery Cc: Microscopy-at-Sparc5.Microscopy.Com Betreff: Re: NO NO NO pellets for Pb stain Datum: Donnerstag, 17. April 1997 01:10
----------------------------------------------------------------------- - The Microscopy ListServer -- Sponsor: The Microscopy Society of America -----------------------------------------------------------------------
} To All } I have been doing TEM since 1970, and have always used NaOH pellets } (CP grade - which do contain about 0.5 - 1% Na2CO3). I make up a 10N } solution in small aliquots (~20 ml), and dump it when I see crystals } of sodium silicate (NaOH dissolves glass). I use this 10N solution } to add to the powdered Pb citrate in dH2O. I make up Pb citrate in } 100 ml quantities and dump it when it turns cloudy. Usually it keeps } for months, unless someone forgets to tighten the cap, which is not } uncommon in a central service lab. What I have learned in nearly 30 } years of TEM, and many years in other microscopy techniques is that } there are very few empirical rules. I have been in very fussy labs } that used aged Pb citrate in bottles that were so coated with } precipitate that they looked out of a sunken pirate ship, and have } been in other labs that make up Pb citrate fresh, and filter it. } Whatever works. } Bruce Cutler, Microscopy Lab, Univ. Kansas, Lawrence KS
I store our freshly made-up concentrated Reynolds solution in 3ml aliquots in Eppendorff tubes at 4C. This way there is minimal exposure to CO2 and no danger of someone leaving the container open. A tube of stain is only used once, anything left over in the tube being discarded. The stain keeps well for months in these tubes.
Robin H Cross Director : EM Unit, Rhodes University, Grahamstown, South Africa eurc-at-giraffe.ru.ac.za - tel: +27 461 318168 - fax: +27 461 24377
Dear colleagues, Does anybody know why everybody uses for section constrasting uranyl acetate which solubility is not so high but not uranyl nitrate or uranyl sulfate which are much more soluble?
Sincerely yours, Alexander Mironov
Consorzio Mario Negri Sud S. Maria Imbaro (Chieti) Italy
Keith: I can't help you with sonicating water, but there is only one thing to do with flat beer, bin it. But then you people in the West Country have no idea what constitutes good beer. Come to Cambridge for a pint of Greene King Abbot or, better still Adnams. Both will knock your socks off
PatrickOn Wed, 16 Apr 1997, Keith Ryan wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Ian } } I really appreciate the tip about sonicating water to degas it, I would not } have believed it. Now, do you have any suggestions for flat beer? } } Regards - Keith Ryan } Plymouth Marine Lab., UK } } }
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America If my memory serves I got sonication from the Technical Hints and Tips in the Proceedings of the RMS. I cover my options by sonicating the water whether fresh or hours, days old. Ian.
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America If my memory serves I got sonication from the Technical Hints and Tips in the Proceedings of the RMS. I cover my options by sonicating the water whether fresh or hours, days old. Ian.
The discussion about UALC staining is wonderful. It's interesting that what works for one doesn't somewhere else.
We went back to Reynold's after problems with the Venable & Coggeshall version. But we store in in 20ml syringes in the refrigerator. It keeps a long time (6-12 months) with no exposure to the air. It filtered through a .22 µm syringe filter. We leave a needle on it and insert it into a rubber stopper. Before use we express a few drops and then place drop onto parafilm in a petri dish.
Good day,
Chuck ------------------------------------- Name: Charles Gilbert VOC:(704)355-5261 Carolinas Medical Center FAX:(704)355-8424 Dept of Pediatric Research PO Box 32861 Charlotte, NC 28232-2861
Chris,
We use 3% glut in 0.1M cacodylate for hematology samples. The membranes are not very well preserved. They are often blurred, partial and sometimes absent but nothing like what you have described. The cells still appear bounded.
We have used cryofixation-freeze substitution and have found excellent membrane preservation in cell suspensions right down to the trilaminar plasmalemma.
Next best is a dimethyl sulfoxide pretreatment before fixing in glut. We use 10% in RPMI-1640 culture media for 10 min. You could probably use whatever media your cells are growing in. This has given us better membrane boundaries than glut alone. They show up unilaminar.
Caveat -- DMSO has some interesting properties and effects on cells. It's best to study the literature especially the safety requirements. It's probably not good to just dump it down the sink.
Best wishes,
Chuck ------------------------------------- Name: Charles Gilbert VOC:(704)355-5261 Carolinas Medical Center FAX:(704)355-8424 Dept of Pediatric Research PO Box 32861 Charlotte, NC 28232-2861
could you please add me to your mailing list (Martin Roe)
In response to Mel Dickson's posting, and to reassure the many Kevex customers on this forum, Kevex is indeed alive and well, but we are experiencing a problem with our Internet service (web site and email) that was just discovered today. As a result, this morning we have been absolutely flooded with telephone calls and other messages reporting this problem. We are sorry for this inconvenience and we are working with our Internet service provider to restore our presence on the Internet as quickly as possible.
Alternate contact methods:
Kevex main office
Tel: +1 (800) 865-3839 (toll free in the USA) Tel: +1 (805) 295-0019 Fax: +1 (805) 295-0419
Kevex main service office
Tel: +1 (800) 495-3839 (toll free in the USA) Tel: +1 (415) 562-2500 Fax: +1 (415) 562-2505
For information on regional offices and other offices worldwide, please contact either of the offices listed above.
Thanks to all who responded to my query about a source for parts. I had thought that Leica was the place and since they are just down the road from us...
In response to Mel Dickson's posting, and to reassure the many Kevex customers on this forum, Kevex is indeed alive and well, but we are experiencing a problem with our Internet service (web site and email) that was just discovered today. As a result, this morning we have been absolutely flooded with telephone calls and other messages reporting this problem. We are sorry for this inconvenience and we are working with our Internet service provider to restore our presence on the Internet as quickly as possible.
Alternate contact methods:
Kevex main office
Tel: +1 (800) 865-3839 (toll free in the USA) Tel: +1 (805) 295-0019 Fax: +1 (805) 295-0419
Kevex main service office
Tel: +1 (800) 495-3839 (toll free in the USA) Tel: +1 (415) 562-2500 Fax: +1 (415) 562-2505
For information on regional offices and other offices worldwide, please contact either of the offices listed above.
I saw their site just a week ago, but am not able to see it today. Maybe something did happen administratively. Anyone else know?
At 01:37 PM 4/17/97 +1000, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
The discussion about UALC staining is wonderful. It's interesting that what works for one doesn't somewhere else.
We went back to Reynold's after problems with the Venable & Coggeshall version. But we store in in 20ml syringes in the refrigerator. It keeps a long time (6-12 months) with no exposure to the air. It filtered through a .22 µm syringe filter. We leave a needle on it and insert it into a rubber stopper. Before use we express a few drops and then place drop onto parafilm in a petri dish.
Good day,
Chuck ------------------------------------- Name: Charles Gilbert VOC:(704)355-5261 Carolinas Medical Center FAX:(704)355-8424 Dept of Pediatric Research PO Box 32861 Charlotte, NC 28232-2861
Chris,
We use 3% glut in 0.1M cacodylate for hematology samples. The membranes are not very well preserved. They are often blurred, partial and sometimes absent but nothing like what you have described. The cells still appear bounded.
We have used cryofixation-freeze substitution and have found excellent membrane preservation in cell suspensions right down to the trilaminar plasmalemma.
Next best is a dimethyl sulfoxide pretreatment before fixing in glut. We use 10% in RPMI-1640 culture media for 10 min. You could probably use whatever media your cells are growing in. This has given us better membrane
boundaries than glut alone. They show up unilaminar.
Caveat -- DMSO has some interesting properties and effects on cells. It's best to study the literature especially the safety requirements. It's probably not good to just dump it down the sink.
Best wishes,
Chuck ------------------------------------- Name: Charles Gilbert VOC:(704)355-5261 Carolinas Medical Center FAX:(704)355-8424 Dept of Pediatric Research PO Box 32861 Charlotte, NC 28232-2861
------------------------------------------------------------------------ The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Reply to: RE} FWD} What has happened to Kevex?
Kevex has a 800 number for field service and messages to others. Try 1-800-495-3839
--------------------------------------
Hello world.
I and other collegues have been trying fruitlessly to e-mail Kevex and when I tried to find their web page it was not there either. Is this just a faulty computer or have they been restructured out of existence? Does anyone out there know?
How do you sonicate to remove gas bubbles? Do you immerse a probe, or place the container in an ultrasonic bath?
Thanks, Jim Williams Indiana University
} } Ultrasonication has been a standard technique for degassing fluids in our } labs for many years. (Before that we boiled where possible or pulled a } gentle vacuum on a closed container. Main purpose was to degas solutions } to be passed through automatic light blockage partcle counters. (Air } bubbles count very nicely as particles.) Nice thing about sonication is } that you can safely sonicate oils and other fluids that you wouldn't want } to boil or pull into a vacuum system. } } Bob Holthausen } Pall Corporation } } } } } } {snip} } If my memory serves I got sonication from the Technical Hints and } Tips in the Proceedings of the RMS. I cover my options by sonicating the } water whether fresh or hours, days old. } Ian.
------------------------------------------------------------------------ The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Reply to: RE} FWD} What has happened to Kevex?
Kevex has a 800 number for field service and messages to others. Try 1-800-495-3839
--------------------------------------
Hello world.
I and other collegues have been trying fruitlessly to e-mail Kevex and when I tried to find their web page it was not there either. Is this just a faulty computer or have they been restructured out of existence? Does anyone out there know?
---------------------------------------------------------- Warning: AOL4FREE.COM is a Trojan Horse Program, which will erase your Hard Disk. ----------------------------------------------------------
Colleagues...
I have checked this announcement out on the U.S. DoE Computer Advisory WWW Page. It is for real so please take notice if you are a DOS/Windows User. Extended details can be found at the WWW address.
http://ciac.llnl.gov/
Technically the program is a Trojan Horse not a virus so it will NOT BE DETECTED by typical virus protection software.
Note, this is a different program from the MacIntosh AOL4FREE program which also circulated the net recently. That one created fraudulant accounts on AOL which is a different issue all together. It was illegal but did not damage your computer. This one will wipe out your disk. A few details are listed below but check out the WWW page at CIAC for the latest information.
A Trojan horse program called AOL4FREE.COM is circulating on the Internet. This is not a virus and cannot be detected by most anti-virus programs. The program is executed by the individual user and deletes all files on a hard drive.
PLATFORM: DOS/Windows-based PCs
DAMAGE: When the AOL4FREE.COM program is executed, all files and directories on the user's C: drive are deleted.
DO NOT execute this program. [Note: Double clicking on.com or .exe will start a selected program.] If the program starts executing, quickly pressing Ctrl-C will save some of your files. If this has happened, shut down immediately and call for assistance. DO NOT attempt to write anything to your hard drive. Files that have been destroyed may be able to be recovered IF you have not written anything to the hard drive and IF your system has not written anything such as when utilizing a auto-save program.
Please see CIAC Bulletin H-47 available on the CIAC homepage (http://ciac.llnl.gov/) for detailed information.
We use 3% glut in 0.1M cacodylate for hematology samples. The membranes are not very well preserved. They are often blurred, partial and sometimes absent but nothing like what you have described. The cells still appear bounded.
We have used cryofixation-freeze substitution and have found excellent membrane preservation in cell suspensions right down to the trilaminar plasmalemma.
Next best is a dimethyl sulfoxide pretreatment before fixing in glut. We use 10% in RPMI-1640 culture media for 10 min. You could probably use whatever media your cells are growing in. This has given us better membrane
boundaries than glut alone. They show up unilaminar.
Caveat -- DMSO has some interesting properties and effects on cells. It's best to study the literature especially the safety requirements. It's probably not good to just dump it down the sink.
Best wishes,
Chuck ------------------------------------- Name: Charles Gilbert VOC:(704)355-5261 Carolinas Medical Center FAX:(704)355-8424 Dept of Pediatric Research PO Box 32861 Charlotte, NC 28232-2861
Research Associate Position
"ELECTRON MICROSCOPY OF MATERIALS"
The Department of Metallurgy and Materials Science, University of Toronto plans to make a term appointment of up to 6 years duration in the area of electron microscopy of materials, with a starting date as early as 1 July 1997. The starting annual salary will be $40,000 plus fringe benefits. The ideal candidate will possess a Ph.D. in a relevant field with significant experience in electron microscopy-based research. The Department operates a facility with a conventional 200 kV TEM/STEM/EDX and 4 computer networked SEM instruments including a high-resolution FESEM with BSE/EDX/EBIC/OIM capabilities. Secondly, a coordinated McMaster University/University of Toronto high-resolution JEOL 200 kV FETEM/STEM with EDX/PEELS/HAADF facility is located at nearby McMaster University. In addition the University of Toronto supports other accessible electron microscopy equipment including a VG HB601 STEM with EDX/PEELS. The candidate will be expected to supervise various activities in the Departmental facility with the support of 3 technical staff members. It is expected that the candidate will be involved in various research projects in conjunction with university/industry users of the facility. The ideal candidate will have experience with a range of microscopy techniques to support ongoing research projects in such areas as semiconductor nanostructures, interfacial segregation analysis, shape memory alloys and biomaterials. Applications, including a curriculum vitae and three letters of reference should be sent to Professor D.D. Perovic, Director, Electron Microscopy Facility, Department of Metallurgy and Materials Science, University of Toronto, 184 College Street, Toronto M5S 3E4 Canada; Fax: (416) 978-4155, Email: perovic-at-ecf.utoronto.ca. The deadline for the receipt of applications is 1 June 1997
_________________ D.D. Perovic Department of Metallurgy and Materials Science, University of Toronto 184 College Street, Toronto M5S 3E4 Canada Tel: (416) 978-5635 Fax: (416) 978-4155
} } } Has anyone comments on the UV polimeriztion of Lowicryl HM20 after freeze } substitution in Acetone-Uranylacetate. I have problems with } prepolymerization of the resin which causes unusable blocks, but the } problem is not consistent, some blocks are good some are not. } } TIA, } Stefan }
I have some experience of this technique. Because HM20 is polymerised by UV light, it is essential that the UV can penetrate the specimen fully, therefore any heavy metals such as osmium or uranyl acetate can potentially cause problems by "shading" areas of the resin from proper polymerisation.
You can minimise the problem by only using mild concentrations of UA in the acetone, by making the blocks as small as possible, and perhaps rinse with pure acetone several times if your substitution apparatus allows it, before polymerising the blocks. Also extend polymerisation time, although this may affect antigenicity. Good luck!
Dare Chris I had the same problem once when I was processing some cultured cells for EM. Morphology was bad at all, but there was no contrast on all membrane. Then I processed the same cell culture with the same batch of osmium solution and fresh osmium side by side. It turned out fresh osmium solved problem.
I have DT2803 frame grabber from Data Translation but without any manual and software. From Data Translation I can't get any information, because this product is out of the production. Please help me. -- Henrik Kaker SEM-EDS Laboratory, Metal Ravne d.o.o., Slovenia Tel: +386-602-21-131, Fax: +386-602-20-436, E-mail: Henrik.Kaker-at-guest.arnes.si http://www2.arnes.si/guest/sgszmera1/index.html Microscopy Vendors DataBase: http://www.kaker.com/mvd/vendors.html Kaker.Com: http://www.kaker.com
Thanks to all of you who responded to the SEM for sale at Mercer University. The microscope has been sold. The folks there are grateful to this forum. ******************************************************* G.W. Erdos, Ph.D. Phone: 352-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax: 352-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 http://www.biotech.ufl.edu/~emcl/ Home of the #1 Gators ***** "Many shall run to and fro, and knowledge shall be increased" Daniel 12:4
} But as an alternative to sonification, you can try just plain BOILING.
Actually, heating to somewhat below boiling will allow the gas to escape. Another possibility is to attach a side-arm flask to the house vacuum and swirl the solution. That works for the case that the solution to be degassed cannot be heated. Yours, Bill Tivol
Ultrasonication has been a standard technique for degassing fluids in our labs for many years. (Before that we boiled where possible or pulled a gentle vacuum on a closed container. Main purpose was to degas solutions to be passed through automatic light blockage partcle counters. (Air bubbles count very nicely as particles.) Nice thing about sonication is that you can safely sonicate oils and other fluids that you wouldn't want to boil or pull into a vacuum system.
Bob Holthausen Pall Corporation
} } On Wed, 16 Apr 1997, Ian Montgomery wrote: } } } Like Rob I've been making Reynold's lead citrate since I was a boy. } } I use NaOH pellets but degas the distilled water by sonicating it for a few } } minutes before making the stain. } } } } Ian. } } Leo, If my memory serves I got sonication from the Technical Hints and Tips in the Proceedings of the RMS. I cover my options by sonicating the water whether fresh or hours, days old. Ian.
Does anyone know the Biosupplies address (Melbourne, Australia)? I want to buy 1,3 , 1,4-B-glucan antibody. Thanks in advance. Nuria Cortadellas University of Barcelona
This is an URGENT request for your comments, suggestions, etc about the 3RD RFD for my proposed new immunocytochemistry newsgroup "sci.bio.immunocytochem" now posted in "news.groups" The latter contains material of a rather varied nature (!) but it is necessary to use this unmoderated group to hold the discussions about all the different proposed new groups.
So, if you are connected to Usenet, and you are keen to see a new immunocytochemistry newsgroup, PLEASE go to "news.groups", ignore all the rubbish, look for articles posted on 16.4.97, and you should find my "3RD RFD: sci.bio.immunocytochem" If your not sure how to post articles to Usenet, select "follow-up article" (or equivalent) from your newsreader menu, and post your message so that it appears under the 3RD RFD.
It is VITALALLY IMPORTANT that some discussion takes place in news.groups soon. Although at least 40 people have e-mailed me to say they want the group to happen (THANK-YOU), but only ONE person has posted any response to my RFDs in "news.groups" where the set-up discussion is supposed to take place.
I am concerned that the lack of discussion indicates a lack of interest in the proposed group, and this is why I have postponed holding the vote. But on the 1st May (election day!) I shall be posting my "CALL FOR VOTES" to the people in charge of Usenet newsgroups. A few days after that, you will be able to vote for the proposed new immunocytochem group.
Amanda Wilson Deputy Manager, E.M. Unit St George's Hospital Medical School, S.W.London, UK Tel: 0181 725 5220 (work) e-mail {awilson-at-aw.u-net.com}
This is an URGENT request for your comments, suggestions, etc about the 3RD RFD for my proposed new immunocytochemistry newsgroup "sci.bio.immunocytochem" now posted in "news.groups" The latter contains material of a rather varied nature (!) but it is necessary to use this unmoderated group to hold the discussions about all the different proposed new groups.
So, if you are connected to Usenet, and you are keen to see a new immunocytochemistry newsgroup, PLEASE go to "news.groups", ignore all the rubbish, look for articles posted on 16.4.97, and you should find my "3RD RFD: sci.bio.immunocytochem" If your not sure how to post articles to Usenet, select "follow-up article" (or equivalent) from your newsreader menu, and post your message so that it appears under the 3RD RFD.
It is VITALALLY IMPORTANT that some discussion takes place in news.groups soon. Although at least 40 people have e-mailed me to say they want the group to happen (THANK-YOU), but only ONE person has posted any response to my RFDs in "news.groups" where the set-up discussion is supposed to take place.
I am concerned that the lack of discussion indicates a lack of interest in the proposed group, and this is why I have postponed holding the vote. But on the 1st May (election day!) I shall be posting my "CALL FOR VOTES" to the people in charge of Usenet newsgroups. A few days after that, you will be able to vote for the proposed new immunocytochem group.
Amanda Wilson Deputy Manager, E.M. Unit St George's Hospital Medical School, S.W.London, UK Tel: 0181 725 5220 (work) e-mail {awilson-at-aw.u-net.com}
Please could you publish the following message on your site, in place of the previous "2nd RFD" Thanks!
PROPOSED NEW IMMUNOCYTOCHEMISTRY NEWSGROUP.......UPDATE!
This is an URGENT request for your comments, suggestions, etc about the 3RD RFD for my proposed new immunocytochemistry newsgroup "sci.bio.immunocytochem" now posted in "news.groups" The latter contains material of a rather varied nature (!) but it is necessary to use this unmoderated group to hold the discussions about all the different proposed new groups.
So, if you are connected to Usenet, and you are keen to see a new immunocytochemistry newsgroup, PLEASE go to "news.groups", ignore all the rubbish, look for articles posted on 16.4.97, and you should find my "3RD RFD: sci.bio.immunocytochem" If your not sure how to post articles to Usenet, select "follow-up article" (or equivalent) from your newsreader menu, and post your message so that it appears under the 3RD RFD.
It is VITALALLY IMPORTANT that some discussion takes place in news.groups soon. Although at least 40 people have e-mailed me to say they want the group to happen (THANK-YOU), but only ONE person has posted any response to my RFDs in "news.groups" where the set-up discussion is supposed to take place.
I am concerned that the lack of discussion indicates a lack of interest in the proposed group, and this is why I have postponed holding the vote. But on the 1st May (general election day here in the UK) I shall be posting my "CALL FOR VOTES" to the people in charge of Usenet newsgroups. A few days after that, you will be able to vote for the proposed new immunocytochem group. I need at least 100 YES VOTES for the group to become official. Amanda Wilson Deputy Manager, E.M. Unit St George's Hospital Medical School, S.W.London, UK Tel: 0181 725 5220 (work) e-mail {awilson-at-aw.u-net.com}
The Russian LOMO scopes have a very good (read low) price. Being somewhat leery of Russian quality, has anyone had any experience with them? They have a Physician/Student model for $650.
Mel Dickson wrote: =================================================== I and other collegues have been trying fruitlessly to e-mail Kevex and when I tried to find their web page it was not there either. Is this just a faulty computer or have they been restructured out of existence? Does anyone out there know? =================================================== Try this FAX #: 1-(805)-295-0419.
Kevex is very much alive and well. Two of their top design and manufacturing people stopped by for their annual visit at our exhibit booth at PITTCON 97 a few weeks ago in Atlanta, and they said they were "very busy" and "working hard to keep up". And coming from them, I would expect that was probably 100% correct!
Chuck
=================================================== Charles A. Garber, Ph. D. Ph: 1-(610)-436-5400 President 1-(800)-2424-SPI SPI SUPPLIES FAX: 1-(610)-436-5755 PO BOX 656 e-mail: cgarber-at-2spi.com West Chester, PA 19381-0656 USA Cust. Service: spi2spi-at-2spi.com
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I have a LOMO low end scope, with added dark field. I am quite pleased with it. It is not as nice to use as a German or Japanese scope, but the optics are quite nice, and you get more of the old fashioned brass, etc. I bought it to use with the kids at home, so I couldn't say how it stands up to industrial use. Built solidly, somewhat homely in paint and finish, but as an optical engineer I am happy with the images.
best regards mark
Mark W. Lund, PhD Director } } Soft X-ray Web page http://www.moxtek.com { { MOXTEK, Inc. 452 West 1260 North Orem UT 84057 801-225-0930 FAX 801-221-1121 lundm-at-xray.byu.edu
"The state is good at simple tasks, like killing people and seizing their wealth. It has far more trouble reaching inside individuals and making them good." Doug Bandow
} I believe that Dr. Mick's interpretation (that REM is german for SEM) is } correct in this case, but there is a technique of Reflection Electron } Microscopy (also abbreviated REM) which is very good for picking out atomic } step edges. See for example: } ... snips
Try also
Reflection Electron Microscopy & Spectroscopy for Surface Analysis Zhong Lin Wang Cambridge University Press ISBN 0 521 48266 6
--------------------------------------------------------------- Dr L. P. Stoter Technical Editor, MICROSCOPY & ANALYSIS Technesis 17, Rocks Park Road email: LPS-at-teknesis.demon.co.uk Uckfield, E. Sussex Phone: +44 (0)1825 766911 TN22 2AT Fax: +44 (0)1825 766911 United Kingdom ---------------------------------------------------------------
I am seeking details of a paper from the 1990 meeting in Seattle. What I have is:
Nicholas G, Bassot J-M, Nicholas M-T (1990). The advantages of cryotechniques: Application to bioluminescent cells. Proceedings of the 12th International Congress of Electron Microscopy, Seattle.
Until we know the details, we can't try for reprint/copy.
This prompts me to ask the obvious question. Does anyone have a foolproof indicator to check the activity of an osmium solution, ideally without processing tissue and viewing in the microscope?
I 'm sure at some time we have all picked up a bottle of expensive osmium and thought do I use it or make fresh up. This happened to me recently and I tried soaking some osmium into a piece of cocktail stick, which appeared to darken, but I was wrong.
Malcolm Haswell University of Sunderland UK ----------
Dare Chris I had the same problem once when I was processing some cultured cells for EM. Morphology was bad at all, but there was no contrast on all membrane. Then I processed the same cell culture with the same batch of osmium solution and fresh osmium side by side. It turned out fresh osmium solved problem.
} I am seeking details of a paper from the 1990 meeting in Seattle. What I } have is: } } Nicholas G, Bassot J-M, Nicholas M-T (1990). The advantages of } cryotechniques: Application to bioluminescent cells. Proceedings of the } 12th International Congress of Electron Microscopy, Seattle. } } Until we know the details, we can't try for reprint/copy
Keith: This was a joint meeting with EMSA and the microbeam society. The proceedings of the meeting published as a four volume set. G.W. Bailey was the publications manager and the San Francisco Press, Inc. published the four volumes. I have Volume 3: Biological Sciences from that meeting. Volume1: Imaging Sciences, Volume 2: Analytical Sciences and Volume 4: Materials Science comprise the titles of the other three volumes. The Nicholas paper was not in volume 3.
The ICEM Program chairs for the meeting were Lee Peachey and D. B. Williams. Good hunting.
Blystone in Texas
-------------------------------- Robert V. Blystone, Ph.D. rblyston-at-trinity.edu
Department of Biology Trinity University 715 Stadium Drive San Antonio, Texas 78212 210.736-7243 FAX 210/736-7229
Dear Colleagues, Does anybody know why everybody uses for staining of ultra thin sections aranyl acetate which has the limited solubility but not uranyl nitrate or uranyl sulfate with the higher solubility.
Sincerely yours, Alexander Mironov
Unit of Morphology Consorzio Mario Negri Sud Via Nazionale, 66030 S. Maria Imbaro (Chieti) Italy
I am a new member on this mailing list. I am receiving replies to the messages that did not submit. And some messages are posted on my mail two or three times. Is this normal?
} I am seeking details of a paper from the 1990 meeting in Seattle. What I } have is: } } Nicholas G, Bassot J-M, Nicholas M-T (1990). The advantages of } cryotechniques: Application to bioluminescent cells. Proceedings of the } 12th International Congress of Electron Microscopy, Seattle. } } Until we know the details, we can't try for reprint/copy
Dear Keith,
I have all five of the volumes from the Seattle meeting. I looked through all of them and I could not find any papers written by Nicholas or Bassot. Are you sure you have the correct meeting?
Good Luck ,
Margaret E. Bisher
NEC Research Institute 4 Independence Way Princeton, NJ 08540. Tel.: (609) 951-2629 Fax: (609) 951-2496 e-mail: peggy-at-research.nj.nec.com
Dear List Is there a reference or does someone have experience with double staining of Epoxy semi-thin sections. I would like to use Basic Fuchsin for red and Toluidine Blue for the blue but I haven't been able to find out any information. Anyone willing to share first hand experience would be appreciated. By the way I am looking for areas with Epstein-Barr viral inclusions in the nucleus. TIA Jack Megill BMS Princeton, NJ
The Nicolas, Bassot, Nicolas talk was the invited lead-off paper in the Cryo-Specimen Prepartation Techniques Symposium chaired buy McDowell and Talon on Thursday August 16, 1990.
The abstract should be in the Imaging Sciences volume unless they did not submit one - unlikely but I recall that this did happen with several invited speakers at the behest of session chairs. This illustrates the problems that inflict the world when guidelines are not enforced.
I will check this volume when I get to where my Proceedings are kept.
There are about 2000 volumes out there so that this matter can be resolved.
} I am seeking details of a paper from the 1990 meeting in Seattle. What I } have is: } } Nicholas G, Bassot J-M, Nicholas M-T (1990). The advantages of } cryotechniques: Application to bioluminescent cells. Proceedings of the } 12th International Congress of Electron Microscopy, Seattle. } } Until we know the details, we can't try for reprint/copy
Dear Keith,
I made a mistake. After seeing Robert Fisher's e-mail I double checked just what I had looked at and I had been looking in the proceedings from the meeting in Paris, Oops!
I did look in the Seattle proceedings and indeed it is there. It is on pages 486 & 487 of Volume 1, Imaging Sciences, just like Dr. Fisher said. If you would like a copy I would be glad to fax it to you or post it in the mail.
Just let me know.
Margaret E. Bisher
NEC Research Institute 4 Independence Way Princeton, NJ 08540. Tel.: (609) 951-2629 Fax: (609) 951-2496 e-mail: peggy-at-research.nj.nec.com
John Megill, Many years ago in graduate school I used to use several different methylene blue-basic fuchsin stains. I remember a special handout provided by LKB that compared these references and showed colored micrographs. They were beautiful. Good Luck!!!! Nancy Monteiro-Riviere
References as follows:
Aparicio, SR, and Marsden, P: Methylene blue-basic fuchsin. Journal of Microsc. (Eng.) 89:139-141, 1969.
Huber, JD, Parker, F, and Odland, GF; Basic fuchsin and alkalinized methylene blue. Stain Technol. 43:83-87, 1968.
Humphrey, CD, and Pittman, Fe: Methylene blue-azure II and basic fuchsin. Stain Technol 42:9-14, 1974.
Nancy A. Monteiro-Riviere, Ph.D.,BCFE,BCFM Professor of Investigative Dermatology/Toxicology North Carolina State University College of Veterinary Medicine Cutaneous Pharmacology and Toxicology Center 4700 Hillsborough Street Raleigh, NC 27606 Telephone: 919-829-4426 FAX: 919-829-4358 email: Nancy_Monteiro-at-ncsu.edu CTPC Homepage: http://cptc.ncsu.edu
We sonicate whatever volume we will need to use in a standard ultrasonic bath. We place solution in a clean beaker and fill the bath with water. Place beaker in the water bath and sonicate for ~15 minutes. If the fluid has a very high viscosity we would sonicate a little longer, maybe 30 minutes.
Bob Holthausen Pall Corporation Scientific and Laboratory Services
williams-at-anatomy.iupui.edu (James C. Williams Jr.) on 04/17/97 03:59:28 PM
To: Microscopy-at-sparc5.microscopy.com -at- internet cc: (bcc: Bob Holthausen/SLSNY/Pall/US)
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} I am seeking details of a paper from the 1990 meeting in Seattle. What I } have is: } } Nicholas G, Bassot J-M, Nicholas M-T (1990). The advantages of } cryotechniques: Application to bioluminescent cells. Proceedings of the } 12th International Congress of Electron Microscopy, Seattle. } } Until we know the details, we can't try for reprint/copy
Dear Keith,
I made a mistake. After seeing Robert Fisher's e-mail I double checked just what I had looked at and I had been looking in the proceedings from the meeting in Paris, Oops!
I did look in the Seattle proceedings and indeed it is there. It is on pages 486 & 487 of Volume 1, Imaging Sciences, just like Dr. Fisher said. If you would like a copy I would be glad to fax it to you or post it in the mail.
Just let me know.
Margaret E. Bisher
NEC Research Institute 4 Independence Way Princeton, NJ 08540. Tel.: (609) 951-2629 Fax: (609) 951-2496 e-mail: peggy-at-research.nj.nec.com
------------------------------------------------------------------------ The Microscopy ListServer -- Sponsor: The Microscopy Society of America
} I am seeking details of a paper from the 1990 meeting in Seattle. What I } have is: } } Nicholas G, Bassot J-M, Nicholas M-T (1990). The advantages of } cryotechniques: Application to bioluminescent cells. Proceedings of the } 12th International Congress of Electron Microscopy, Seattle. } } Until we know the details, we can't try for reprint/copy
Dear Keith,
I made a mistake. After seeing Robert Fisher's e-mail I double checked just what I had looked at and I had been looking in the proceedings from the meeting in Paris, Oops!
I did look in the Seattle proceedings and indeed it is there. It is on pages 486 & 487 of Volume 1, Imaging Sciences, just like Dr. Fisher said. If you would like a copy I would be glad to fax it to you or post it in the mail.
Just let me know.
Margaret E. Bisher
NEC Research Institute 4 Independence Way Princeton, NJ 08540. Tel.: (609) 951-2629 Fax: (609) 951-2496 e-mail: peggy-at-research.nj.nec.com
This is a protocol we have used that gives wonderful results:
Polychrome stain
1. Blue Stain: Methylene blue .13gm , Azure II .02gm, Glycerol 10ml, Methyl alcohol 10ml, D.H2O 80ml (stir and filter, keeps 6 mo.)
2. Red stain: STOCK SOL: Basic fuchsin .2gm, DH2O 100ml (stir and filter, keeps 6mo.) WORKING SOL: dilute stock 1:4 in DH2O (fresh daily)
3. Sodium Hydroxide 1% fresh daily
Procedure: 1. Flood slide with blue stain 15-60 seconds, depending on temperature and material. 2. Add 4-6 drops NaOH to the stain and mix by tilting the slide, about 10 seconds total time. 3. Wash in running water and dry on hotplate. Blue stain can be destained by heating. 4. Add red stain for 15-30 seconds on hotplate. (Red stain cannot be destained) 5. Rinse with running water and dry.
References: Mackay and Mead. 1970. 28th EMSA meetings pp.296-297. Modified by Griffin and Fahrenbach, Oregon Regional Primate Research Center
Hope this is of some help Linda M. Fox Dept. of CBN and Anatomy Loyola University Medical School 2160 S. First Ave. Maywood, Illinois 60153 lfox1-at-wpo.it.luc.edu
I am a new member on this mailing list. I am receiving replies to the messages that did not submit. And some messages are posted on my mail two or three times. Is this normal?
Hong Yi
Emory, Neurology
Basically, yes. This message has CC: Microscopy-at-Sparc5.Microscopy.Com so it will be sent to the list. It also has To: hyi-at-emory.edu so it will go straight to you. You will probably receive two copies. Typical mailers will put in the CC: header when a "reply" command is issued, so everyone on the list will get replies unless the sender specifically deleted the CC: header before sending the reply.
Sometimes users send in a message, wait an hour to see if it really went in, and then send it again because they haven't seen it yet. This can cause multiple copies of messages. There are other possible causes as well. I noticed a few duplicates recently but didn't spend the time to figure out how they happened.
I was wondering if anyone knew of a good fix for TEM studies of ciliated protists in marine organisms such as oysters. I've tried different fixes and the tissue looks good but the protists....uhhhh looks bad. Any suggestions? Thanks.
Peace through Christ,
Phil Rutledge, Director e-mail: prutle1-at-gl.umbc.edu Center for Microscopy voice: (410) 455-3582 UMBC Dept. of Biology fax: (410) 455-3875 Catonsville, MD 21228 ///// / / / / /////// /////// / / /////// /////// / / / / / / / / / / / / /////
Received: by dub-img-5.compuserve.com (8.6.10/5.950515) id PAA07822; Fri, 18 Apr 1997 15:07:01 -0400 Comments: Returned from: {74767.3673-at-CompuServe.COM} Message-Type: Delivery Report Message-ID: {970418190028_515664.456256_GHQ43-46-at-CompuServe.COM}
This is a protocol we have used that gives wonderful results:
Polychrome stain
1. Blue Stain: Methylene blue .13gm , Azure II .02gm, Glycerol 10ml, Methyl alcohol 10ml, D.H2O 80ml (stir and filter, keeps 6 mo.)
2. Red stain: STOCK SOL: Basic fuchsin .2gm, DH2O 100ml (stir and filter, keeps 6mo.) WORKING SOL: dilute stock 1:4 in DH2O (fresh daily)
3. Sodium Hydroxide 1% fresh daily
Procedure: 1. Flood slide with blue stain 15-60 seconds, depending on temperature and material. 2. Add 4-6 drops NaOH to the stain and mix by tilting the slide, about 10 seconds total time. 3. Wash in running water and dry on hotplate. Blue stain can be destained by heating. 4. Add red stain for 15-30 seconds on hotplate. (Red stain cannot be destained) 5. Rinse with running water and dry.
References: Mackay and Mead. 1970. 28th EMSA meetings pp.296-297. Modified by Griffin and Fahrenbach, Oregon Regional Primate Research Center
Hope this is of some help Linda M. Fox Dept. of CBN and Anatomy Loyola University Medical School 2160 S. First Ave. Maywood, Illinois 60153 lfox1-at-wpo.it.luc.edu
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Re:
} Nicholas G, Bassot J-M, Nicholas M-T (1990). The advantages of } cryotechniques: Application to bioluminescent cells. Proceedings of the } 12th International Congress of Electron Microscopy, Seattle.
Dear Robert,
Why not write directly to Marie-Therese Nicholas at Laboratoire de Bioluminescence CNRS 105 Blvd Raspail 75006 Paris France
As an author, she should be able to help you.
Regards,
Paul Webster, Ph.D Center for Cell Imaging Yale School fo Medicine http://info.med.yale.edu/cellimg
------------------------------------------------------------------------ The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Re:
} Nicholas G, Bassot J-M, Nicholas M-T (1990). The advantages of } cryotechniques: Application to bioluminescent cells. Proceedings of the } 12th International Congress of Electron Microscopy, Seattle.
Dear Robert,
Why not write directly to Marie-Therese Nicholas at Laboratoire de Bioluminescence CNRS 105 Blvd Raspail 75006 Paris France
As an author, she should be able to help you.
Regards,
Paul Webster, Ph.D Center for Cell Imaging Yale School fo Medicine http://info.med.yale.edu/cellimg
I've had no problems with Reynold's lead citrate, using freshly boiled, distilled water and 1N NaOH freshly prepared from pellets. I store the finished stain in a disposable syringe with all air bubbles expelled, and the tip covered in plastifilm. For use I attach a 0.2 micron syringe filter and needle and expel a few drops first, then use the next drops. As long as there is no air in the syringe, and it is kept in the refrig, it will keep forever? or at least 8 months. The recipe: 1.33 g lead nitrate, 1.76 g sodium citrate mixed with 30 ml freshly boiled distilled or deionized water- mix in a 50 ml volumetric flask- shake vigorously for 1 min and intermittently for 30 mins. Solution will be milky.Add 8.0 ml of 1 N NaOH freshly prepared.Solution becomes clear. Dilute to 50 ml with freshly boiled, distilled or deionized water. Load into syringe- I use a 60 cc disposable distilled or deionized water. Load into syringe- I use a 60 cc disposableI,ve been using this method for 20 years consistent success. Just my two-cents worth.
Pat Masarachia Bone Biology Merck and Co. West Point, PA 215-652-7999
So why do mail problems happen on Friday Afternoons?
Maybe that 's when accounts are deleted or something. I've been doing some investigation on this afternoons set of problems. There was an intermittent fault in mail duplicaton. If your interested read on, otherwise delete this and just consider this as just a test message.
The duplicate transmission was caused by the mail nameserver hanging on a memory/queue failure. This was due to a number of sites (~ 10) either going down and/or just going out of existance (host name unkown) plus a number of user accounts either filling (mailbox full) or being deleted (user unknown) without the user unsubscribing (~20) . The net effect of all these was that the queue of undelivered messages started to grow significantly. This in turn started a new set of delivery queuese which began to eat up even more CPU resources.
Eventually a few of the queues failed and but when message queue fails the sendmail program tries to resend the entire message, however, it (sendmail program) forgot that it had sent the message to part of the mailing list already and so a large number of people got multiple copies, depending upon where you name feel in the subscription list.
I've tried to reconfigure the server to minimize this happening again, however, to do so I've instituted yet another queuing system for all listserver mail. Instead of trying to immediately deliver all mail as we did in the past, everything is now put into a small queue and delayed ~ 15-30 minutes before starting a delivery sequence.
This does not mean that you will see a message 15-30 minutes after someone submits it, but the process of running the mail server becomes more regimented.
Due to the size of the mailing list and the sometime poor links to some sites mail can take more than an hour (or longer) to propagate through the entire list. I've also attempted to setup a mail configuration file to minimize duplicate deliveries in the case of a queue failure. If things work correctly no more than 10 people should receive duplicate messages on a crash, as the delivery list is rewritten now every 10 deliveries. This will also (obviously) increase the time it takes to process the mail.
Let's see if this cures the duplicate delivery problem... (fingers crossed and blurry eye he reaches for the send button)
G'night all.
Nestor Yawn.. Your tired Friendly Neighborhood SysOp
Q: How many internet mail list subscribers does it take to change a light bulb?
A: 1,331:
1 to change the light bulb and to post to the mail list that the light bulb has been changed. 14 to share similar experiences of changing light bulbs and how the light bulb could have been changed differently. 7 to caution about the dangers of changing light bulbs. 27 to point out spelling/grammar errors in posts about changing light bulbs. 53 to flame the spell checkers. 156 to write to the list administrator complaining about the light bulb discussion and its inappropriateness to this mail list. 41 to correct spelling in the spelling/grammar flames. 109 to post that this list is not about light bulbs and to please take this email exchange to alt.lite.bulb. 203 to demand that cross posting to alt.grammar, alt.spelling and alt.punctuation about changing light bulbs be stopped. 111 to defend the posting to this list saying that we are all use light bulbs and therefore the posts **are** relevant to this mail list. 306 to debate which method of changing light. bulbs is superior, where to buy the best light bulbs, what brand of light bulbs work best for this technique, and what brands are faulty. 27 to post URLs where one can see examples of different light bulbs. 14 to post that the URLs were posted incorrectly, and to post corrected URLs. 3 to post about links they found from the URLs that are relevant to this list which makes light bulbs relevant to this list. 33 to concatenate all posts to date, then quote them including all headers and footers, and then add "Me Too.". 12 to post to the list that they are unsubscribing because they cannot handle the light bulb controversy. 19 to quote the "Me Too's" to say, "Me Three.". 4 to suggest that posters request the light bulb FAQ. 1 to propose new alt.change.lite.bulb newsgroup. 47 to say this is just what alt.physics.cold_fusion was meant for, leave it there.
I've received enough requests for copies of the general information and FAQ messages that I've now put them up on-line for you to access via the WWW at your convenience.
Just go to the MSA WWW home page and follow the Microscopy ListServer Links.
http://www.msa.microscopy.com
I will continue to send out Email versions with every new subscription or to anyone requesting an Emailed copy.
Nestor Your Friendly Neighborhood SysOp.
-------- BTW,... (with fingers crossed) no duplicate messages so far!
I am study artifacts on trasmission electron microscopy. I have not bibliography. Please your send references. Thank you Dr. Alberto Pizarro G. Histopathology rediegal-at-homonet.com.mx
Dear Colleagues, Does anybody know why for the staining of ultra thin sections everybody uses uranyl acetate which has rather limited solubility in water, but not yranyl sulfate or uranyl nitrate which are much more soluble.
Alexander Mironov
Unit of Morphology Consorzio Mario Negri Sud S. Maria Imbaro (Chieti) Italy
} This prompts me to ask the obvious question. Does anyone have a foolproof } indicator to check the activity of an osmium solution, ideally without } processing tissue and viewing in the microscope? } } I 'm sure at some time we have all picked up a bottle of expensive osmium } and thought do I use it or make fresh up. This happened to me recently and I } tried soaking some osmium into a piece of cocktail stick, which appeared to } darken, but I was wrong. } } Malcolm Haswell
Malcolm, I would put a drop of corn oil (maize oil on your side of the puddle) on a slide and some of the osmium solution in a small dish (size so that the slide acts as a lid for the dish). If the osmium is good, vapors from it will blacken the oil droplet. (Any polyunsaturated oil will do.) Phil
&&& Illigitimi non carborundum &&&&&&&& Philip Oshel Station A PO Box 5037 Champaign, IL 61825-5037 (217) 355-1143 oshel-at-ux1.cso.uiuc.edu *** looking for a job again ******************
Seattle paper: thanks to everyone for the help in finding details of the paper in question. It was found and offers made of a copy!
Flat beer: most of this was off line - apologies for that which crept back on-line. The result is I am bearing home brew kits to a new found friend in Chicago next month! So, some good came of it!
Thanks again from Plymouth UK (on a grey Monday morn) Keith Ryan
10 BIT MONOCHROME } 62dB S/N REAL TIME CAMERAS, FRAME GRABBERS, & SOFTWARE INFO / NEW WEB SITE DVC Company is a US manufacturer of video cameras and has a updated web with detailed info /Q & A on: DVC-10 10 bit digital & 10 bit analog monochrome CCD cameras DVC-8 8 bit digital & 10 bit analog monochrome CCD cameras DVC-0A 10 bit analog monochrome camera (upgradable) to digital Complete frame grabber PCI bus board listing supplied by DVC as a systems house, along with imaging software. ((( DVC can offer a complete package ))) camera, digital cable, frame grabber, software, and tuneable monochrome and RGB LCD filters http://members.aol.com/dvcco or http://www.edt.com/dvc/dvc.html Fill in the form for contact and or: Download the complete DVC 80 page manual, data sheet, and TI sensor info via Acrobat 3.0 reader for selective print out later. DVC Company/ San Diego, CA 619-444-8300 619-444-8321-fax This posting is only on two newsgroups due to the focus of the products.
I am mailing you regarding membership to the American Society of Electron Microscopy. Arthur Slaughter and myself are electron microscopists at HRL Technology, Melbourne Australia, and are currently members of the Australian Society for Electron Microscopy. HRL Technology is a Research and Development laboratory which was originally part of the State Electricity Commission of Victoria. The Electron Microscope Facility at HRL is mainly used for solving materials related problems for a wide range of industries in Australia, including mineral and metals processing, mining and power production, building and food industries to name a few.
We are interested in subscribing to the American Society of Electron Microscopy as we believe this would benefit our day to day work, and provide information regarding current areas of concern in the US.
Could you please send us information regarding ASEM, and any conditions required for membership to this organisation.
MAMAS (Mid-Atlantic Microbeam Analysis Society) and the Surface and Microanalysis Science Division, NIST Meetin at the National Institute of Standards and Technology, Gaithersburg, MD on Thursday, May 15, 1997, 10:30 am- 3:00 PM Lecture Room D, Administration Bldg.
10:30am Coffee and Doughnuts
10:45am Prof. David R. Veblen, Dept. of Earth and Planetary Science, Johns Hopkins University "Transmission Electron Microscopy of Minerals"
12 noon Lunch
1:15pm Dr. Michael Kersker, TEM/STM Project Manager, JEOL USA "200 KV FEG: Refried Beans with a Fiery New Salsa"
For more information, contact Ryna Marinenko (301)975-3901, FAX(301)417-1321, email:ryna.marinenko-at-nist.gov
We work as consultants for a computer imaging hardware and software manufacturer's rep, and have had a rather unusual request. Perhaps someone, either end users or vendors can assist.
We are looking for the equivalent of a MacBeth Color Chart, typically used in video, but this needs to be very small, translucent, and mounted on a standard histo glass slide. Ideally, 1/32" square (1 mm x 1 mm OK) and it would contain at least the primary colors, incl. black and white. I suspect there may be a problem with ?translucent black and white. I think episcopic illumination would also suffice for the chart, however so it could be opaque. At this time, I do not have more detail on their exact application.
We have checked with Munsell and MacBeth to no avail, so we would appreciate any assistance, and even a referral as it's likely this is a custom application. You can e-mail me directly if you like.
I am sending this message again because it was returned, in error on Friday. ----------
This prompts me to ask the obvious question. Does anyone have a foolproof indicator to check the activity of an osmium solution, ideally without processing tissue and viewing in the microscope?
I 'm sure at some time we have all picked up a bottle of expensive osmium and thought do I use it or make fresh up. This happened to me recently and I tried soaking some osmium into a piece of cocktail stick, which appeared to darken, but I was wrong.
Malcolm Haswell University of Sunderland UK ----------
Dare Chris I had the same problem once when I was processing some cultured cells for EM. Morphology was bad at all, but there was no contrast on all membrane. Then I processed the same cell culture with the same batch of osmium solution and fresh osmium side by side. It turned out fresh osmium solved problem.
Message-ID: {01BC4E50.DFE665E0-at-pguerin.lisco.com} {CONFOCAL-at-LISTSERV.ACSU.BUFFALO.EDU} , "'MICROSCOPY-at-SPARC5.MICROSCOPY.COM'" {MICROSCOPY-at-Sparc5.Microscopy.Com} Cc: Chris MacLean {Windows/pguerin/cmaclean-at-vaytek.com}
This message is from a software vendor.
Hello,
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Group - I would like to organize a summary listing of all non-supplier/manufacturer web pages of interest to microscopists. I already do the supplier/manufacturer listing. Format to be WWW address, organizers name and establishment and a some-100 max word description of the site. When complete I will publish the full listing on this listserver as well as in my publication. All help would be much appreciated. Don Grimes, Microscopy Today
Maybe y'all can help this soul out until he can get subscribed.
} Return-Path: {muellerd-at-mis.finchcms.edu} } Date: Mon, 21 Apr 1997 14:55:43 -0700 } From: David Mueller {muellerd-at-mis.finchcms.edu} } Reply-To: muellerd-at-mis.finchcms.edu } Organization: The Chicago Medical School } To: sdw-at-biotech.ufl.edu } Subject: collodial gold labeling } } I am uncertain how to access this newsgroup on EM. Could you give me } the } address? } } Alternatively, maybe you know the answer. I need to label MAb with } colloidal gold. Since the Ab is available in only small amount, I } prefer not to optimize the conditions as it requires a lot of Ab. Are } there standard conditions to label MAb with gold? What is the minimal } protein concentration needed to get effective binding? Can BSA be added } to stabilize the binding and fill the unbound sites? } } Thanks for any help in the matter. } } David Mueller } Dept. Biological Chemistry } The Chicago Medical School } muellerd-at-mis.finchcms.edu } }
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { GO GATORS Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 352-392-1295 ICBR EM Core Lab fax 352-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/ The home of " Tips & Tricks "
A colleage of mine, Lonnie Russell, has co-authored an introductory book on molecular biology (MB) that may be of interest to microscopists. Since MB may be new or intimidating (yet is important for microscopists) this might be one book to consider. If you would like more information, he may be contacted directly at {lrussell-at-som.siu.edu} . Note: I have no financial interest in this book - I just find it useful.
#################################################################### John J. Bozzola, Ph.D., Director Center for Electron Microscopy Neckers Building, Room 146 - B Wing Southern Illinois University Carbondale, IL 62901-4402 U.S.A. Phone: 618-453-3730 Fax: 618-453-2665 Email: bozzola-at-siu.edu Web: http://www.siu.edu/departments/shops/cem.html ####################################################################
I am looking into upgrading our present image Analysis system (Kontron IBAS).
Has anyone performed extensive comparisons between the leading IA manufacturers who is willing offer pro's and con's of each IA system they have examined.
Some of the IA systems include (but not limited to)
Kontron KS400 Optimas Mediacybernetics Image Pro Plus ImagePro (Nikon) Bioquant Quantimat Noesis (Visilog5)
Applications include wide variety of biological (Pathology/toxicology applications, density gradients, fluorescence, stereology and morphometry etc.), and non biological (particle size, coating thickness, fiber analysis, phase boundary etc.).
Ease of programming (Macro scripting or interpreter language) and program modification for those who are not computer programers is a must.
Input on camera and video capture boards are welcomed as well.
Thank you in advance for any information you are willing to share.
The heaviest element known to science was recently discovered. The element, tentatively named ADMINISTRATIUM, has no protons or electrons and thus has an atomic number of 0.
However, it does have 1 Neutron, 128 Assistant Neutrons, 75 Vice-Neutrons and 111 Assistant Vice Neutrons. This gives it an atomic weight of 315. These 315 particles are held together in a nucleus by a force that involves the continuous exchange of meson-like particles called Morons.
Since it has no electrons, Administratium is inert. However, it can be detected chemically as it impedes every other reaction with which it comes into contact. According to the discoverers, a minute amount of Administratium caused one reaction to take over four days to complete, when it would normally occur in less than one second.
Administratium has a normal life of approximately 3 years, at which time it does not decay, but instead, undergoes a reorganization in which Assistant Neutrons, Vice-Neutrons and Assistant Vice-Neutrons exchange places. Some studies have shown that the atomic weight actually increases after each reorganization.
Research at other laboratories indicates that Administratium occurs naturally in the atmosphere. It tends to concentrate at certain points such as government, large companies, healthcare facilities and universities; and will often be found in the newest, best maintained buildings.
Scientists point out that Administratium is know to be toxic at any level of concentration and can easily destroy any productive reactions where it is allowed to accumulate.
Sad but all too true !!
Sent to me by
JamesR0712-at-aol.com \\|// (o o) ~~~~~~~~~~~~oOOo~(_)~oOOo~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
{ { {This message is made of 100% recycled electrons} } }
Cheers ;o) :o) %o) Eric {Mesa Arizona} http://www.goodnet.com/~earosen (Note the tilde before earosen)
Do you really need to label your antibody? Rather, I'd recommend that you use an indirect labelling method in which, e.g., the secondary antibody is coupled to gold or is biotinylated and finally detected with streptavidin gold.
If you really need to gold label your MAb, you will find the necessary information and protocols in the reviews and papers published by J. Roth in the late 70's and early 80's, as well as in many books on immunolabelling.
Regards, Michel
**************************************************** Michel Deschuyteneer, Ph.D. deschuyt-at-sbbio.be Scientist Electron Microscopy Laboratory
SmithKline Beecham Biologicals Rue de l'Institut, 89 B1330 Rixensart, BELGIUM Tel: +32-2-656 9290 Fax: +32-2-656 8164 **************************************************** Standard disclaimer: the opinions expressed in this communication are my own and do not necessarily reflect those of SmithKline Beecham. ****************************************************
I am looking for the fax number or email address of Scanning Microscopy Int. (Chicago) - can anybody help?
Thanks in advance Fiona Graham Electron Microscope Unit University of Natal, Dalbridge, 4041, South Africa tel: +27 31 260 2174 fax: +27 31 261 6550 email: GRAHAM-at-ph.und.ac.za URL: http://www.und.ac.za/und/emu/emunit.html
You wrote: " I am looking for the fax number or email address of Scanning Microscopy Int. (Chicago) - can anybody help?
Thanks in advance Fiona Graham Electron Microscope Unit University of Natal, Dalbridge, 4041, South Africa tel: +27 31 260 2174 fax: +27 31 261 6550 email: GRAHAM-at-ph.und.ac.za"
I have used 73211.647-at-compuserve.com within the last couple of months and it was OK.
Donald J. Marshall Relion Industries P.O. Box 12 Bedford, MA 01730 Ph: 617-275-4695 FAX: 617-275-4695 Alternate FAX: 617-271-0252
DMartin/RRosencrans wrote: } } ------------------------------------------------------------------------} The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------.} } Hello all, } } We work as consultants for a computer imaging hardware and software } manufacturer's rep, and have had a rather unusual request. Perhaps someone, } either end users or vendors can assist. } } We are looking for the equivalent of a MacBeth Color Chart, typically used } in video, but this needs to be very small, translucent, and mounted on a } standard histo glass slide. Ideally, 1/32" square (1 mm x 1 mm OK) and it } would contain at least the primary colors, incl. black and white. I suspect } there may be a problem with ?translucent black and white. I think episcopic } illumination would also suffice for the chart, however so it could be } opaque. At this time, I do not have more detail on their exact application. } } We have checked with Munsell and MacBeth to no avail, so we would appreciate } any assistance, and even a referral as it's likely this is a custom } application. You can e-mail me directly if you like. } } Thanks in advance!! } } Daryl Martin } dmartin-at-ic.net } } (313) 213-8444Dear Daryl, When I worked at Zeiss as microspectrophotometry specialist, one of my colleagues made a wonderful test slide using very small strips of colored film, laid side by side on a slide then covered with a coverslip. It sounds like a modification would be ideal for your purposes.
Hope this helps. Barbara Foster Consortium President Microscopy/Marketing & Education
Gregory.Argentieri-at-sandoz.com wrote: } } ------------------------------------------------------------------------} The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------.} } Dear Microscopists: } } I am looking into upgrading our present image Analysis system (Kontron } IBAS). } } Has anyone performed extensive comparisons between the leading IA } manufacturers who is willing offer pro's and con's of each IA system } they have examined. } } Some of the IA systems include (but not limited to) } } Kontron KS400 } Optimas } Mediacybernetics Image Pro Plus } ImagePro (Nikon) } Bioquant } Quantimat } Noesis (Visilog5) } } Applications include wide variety of biological (Pathology/toxicology } applications, density gradients, fluorescence, stereology and } morphometry etc.), and non biological (particle size, coating } thickness, fiber analysis, phase boundary etc.). } } Ease of programming (Macro scripting or interpreter language) and } program modification for those who are not computer programers is a } must. } } Input on camera and video capture boards are welcomed as well. } } Thank you in advance for any information you are willing to share. } } Greg } } Gregory Argentieri } Novartis Pharmaceuticals Corp. } Gregory.Argentieri-at-pharma.novartis.com } Gregory.Argentieri-at-sandoz.com } Greg2NJ-at-aol.com } } 201-503-8617Dear Greg, The list you sent suggests that you may want to clarify your needs a little further before you go shopping. 1. Do you want a fully integrated system or something which is more modular? Quantimat and Kontron systems are sold as systems whereas Media Cy, Optimas, and Noeisis are software only. 2. You mentioned the level of programming. I am more familiar with the off-the-shelf systems and of those, Noesis is the most powerful but requires the greatest sophistication in terms of programming ability. Media Cy's Image Pro Plus and Optimas are more mid-range products. We have conducted a variety of market research projects, both at meetings (Microscopy & MicroAnalysis, ASMaterials, Experimental Bio, etc.), by phone, and personal interview. Of these two packages, Media Cy's Image Pro is the more widely accepted software. Several system integrators also indicated that it is more user friendly and the package has been adopted by a number of the microscope companies (ex: Zeiss' Image One is a privately labeled version; I expect that Nikon's offering is the same, Topcon and Phillips have used IPP in conjunction with their EMs). 3. Re: the hardware - we strongly suggest that you talk to a few local system integrators. They can give you the latest on the hardware/software/camera technologies and put together a system which meets your needs. (Input from other colleagues never hurts, however).
Hope this helps. If you need training when you get your system together, give us a call.
Barbara Foster Consortium President Microscopy/Microscopy Education
We have no financial interest in any of the products mentioned above.
Here is a request I am forwarding for a colleague, Daniel Wilcox. Thank you for your help.
} Audrey Dow } } I am looking for experienced Hitachi 4500-II FESEM users that have used } their instruments for electron beam induced current (EBIC) imaging of } gallium arsendie microwave devices. I am looking for advice on how to } build or buy a sub-stage for the large Hitachi stage (with a 6" } intro-port) that will allow for mechanical probe tips to contact device } circuit elements, and allow DC bias to be applied as well as the induced } current signal to be fed to our GW Electronics speciman current amp. I } have done extensive EBIC with our Cambridge 250 SEM, but the amount of } sample current available in the Hitachi is much less. } } If anyone has some suggestions, please contact me at 301-428-4233 at } M/A-COM, e-mail "wilcoxd-at-macom.com". Thanks! } } Daniel Wilcox
(Since Daniel sometimes has trouble with e-mail, you can send messages to me at audrey.dow-at-amp.com and I will forward them.) }
The printing inks used to create the MacBeth Color Chart cannot be precisely duplicated with transparency materials. It is possible to create a close approximation by carefully photographing a color chart with 35mm transparency film. I have used a Kodachrome slide of the MacBeth chart included with a Beseler slide duplicator for calibration of that system. If you are photographing your own chart the most rigorous results will be obtained if you check the slides with a densitometer and correct for any density shifts or color shifts then rephotograph. Once you have a good 35mm slide then you can reduce the size by carefully rephotographing using the same type of controls as above. Hopefully you can work with a good professional film processing laboratory. Their knowledge of color materials and sensitometry can be quite helpful. Keep in mind that film materials vary in their permanency. Some will fade non-linearly quite rapidly when exposed to intense light levels. Larry D. Ackerman (415) 476-8751 Howard Hughes Medical Institute FAX (415) 476-5774 UCSF, Box 0724, Rm U426 533 Parnassus Ave. mishot-at-itsa.ucsf.edu San Francisco, CA 94143
The Conservation Analytical Laboratory (CAL) of the Smithsonian Institution, located in Suitland, MD, is recruiting for a Microscopist, GS-11 ($38,330-$49,831) or GS-12 ($45,919-59,725). Duties include optical microscopy, chemical microscopy, freezing-heating stage analysis, digital imaging and analysis, sample preparation, and microscope maintenance. The incumbent will be required to actively participate in the educational/training activities of CAL.
At least an undergraduate degree in physical/biological science or related field is required. The applicant must have knowledge of the principles and experience in the practice of optical microscopy, and skill in conducting training in a technical field to a professional audience.
For a copy of the vacancy announcement, call the Smithsonian 24-hour Automated Jobline (202) 287-3102, press 9 and request Announcement # 97PL-3079. Applications must be postmarked by June 18, 1997. The Smithsonian Institution is an Equal Opportunity Employer.
} ------------------------------------------------------------------------ } } The Microscopy ListServer -- Sponsor: The Microscopy Society of } America } To Subscribe/Unsubscribe -- Send Email to } ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } Gregory.Argentieri-at-sandoz.com wrote: } } } } } ------------------------------------------------------------------------} } The Microscopy ListServer -- Sponsor: The Microscopy Society of } America } } To Subscribe/Unsubscribe -- Send Email to } ListServer-at-MSA.Microscopy.Com } } } -----------------------------------------------------------------------.} } } } Dear Microscopists: } } } } I am looking into upgrading our present image Analysis system } (Kontron } } IBAS). } } } } Has anyone performed extensive comparisons between the } leading IA } } manufacturers who is willing offer pro's and con's of each IA } system } } they have examined. } } } } Some of the IA systems include (but not limited to) } } } } Kontron KS400 } } Optimas } } Mediacybernetics Image Pro Plus } } ImagePro (Nikon) } } Bioquant } } Quantimat } } Noesis (Visilog5) } } } } Applications include wide variety of biological } (Pathology/toxicology } } applications, density gradients, fluorescence, stereology and } } } morphometry etc.), and non biological (particle size, coating } } } thickness, fiber analysis, phase boundary etc.). } } } } Ease of programming (Macro scripting or interpreter language) } and } } program modification for those who are not computer } programers is a } } must. } } } } Input on camera and video capture boards are welcomed as } well. } } } } Thank you in advance for any information you are willing to } share. } } } } Greg } } } } Gregory Argentieri } } Novartis Pharmaceuticals Corp. } } Gregory.Argentieri-at-pharma.novartis.com } } Gregory.Argentieri-at-sandoz.com } } Greg2NJ-at-aol.com } } } } 201-503-8617Dear Greg, } The list you sent suggests that you may want to clarify your needs a } } little further before you go shopping. } 1. Do you want a fully integrated system or something which is more } modular? Quantimat and Kontron systems are sold as systems whereas } Media } Cy, Optimas, and Noeisis are software only. } 2. You mentioned the level of programming. I am more familiar with } the } off-the-shelf systems and of those, Noesis is the most powerful but } requires the greatest sophistication in terms of programming } ability. } Media Cy's Image Pro Plus and Optimas are more mid-range products. } We } have conducted a variety of market research projects, both at } meetings } (Microscopy & MicroAnalysis, ASMaterials, Experimental Bio, etc.), } by } phone, and personal interview. Of these two packages, Media Cy's } Image } Pro is the more widely accepted software. Several system } integrators } also indicated that it is more user friendly and the package has } been } adopted by a number of the microscope companies (ex: Zeiss' Image } One is } a privately labeled version; I expect that Nikon's offering is the } same, } Topcon and Phillips have used IPP in conjunction with their EMs). } 3. Re: the hardware - we strongly suggest that you talk to a few } local } system integrators. They can give you the latest on the } hardware/software/camera technologies and put together a system } which } meets your needs. (Input from other colleagues never hurts, } however). } } Hope this helps. If you need training when you get your system } together, } give us a call. } } Barbara Foster } Consortium President } Microscopy/Microscopy Education } } We have no financial interest in any of the products mentioned } above.
Greg - since you are upgrading you might also look at Amerinex's Aphelon software. It seems to be very powerful being a spinoff of some AI software developed for the military. I believe that they are at http://www.amerinex.com. If you are willing to program a system youmight also look at ContextVision's MicroGOP - they are from Sweden and the only number I have is there - 46-13-102480 or FAX 46-13-104282.
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} ------------------------------------------------------------------------ } } The Microscopy ListServer -- Sponsor: The Microscopy Society of } America } To Subscribe/Unsubscribe -- Send Email to } ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } Gregory.Argentieri-at-sandoz.com wrote: } } } } } ------------------------------------------------------------------------} } The Microscopy ListServer -- Sponsor: The Microscopy Society of } America } } To Subscribe/Unsubscribe -- Send Email to } ListServer-at-MSA.Microscopy.Com } } } -----------------------------------------------------------------------.} } } } Dear Microscopists: } } } } I am looking into upgrading our present image Analysis system } (Kontron } } IBAS). } } } } Has anyone performed extensive comparisons between the } leading IA } } manufacturers who is willing offer pro's and con's of each IA } system } } they have examined. } } } } Some of the IA systems include (but not limited to) } } } } Kontron KS400 } } Optimas } } Mediacybernetics Image Pro Plus } } ImagePro (Nikon) } } Bioquant } } Quantimat } } Noesis (Visilog5) } } } } Applications include wide variety of biological } (Pathology/toxicology } } applications, density gradients, fluorescence, stereology and } } } morphometry etc.), and non biological (particle size, coating } } } thickness, fiber analysis, phase boundary etc.). } } } } Ease of programming (Macro scripting or interpreter language) } and } } program modification for those who are not computer } programers is a } } must. } } } } Input on camera and video capture boards are welcomed as } well. } } } } Thank you in advance for any information you are willing to } share. } } } } Greg } } } } Gregory Argentieri } } Novartis Pharmaceuticals Corp. } } Gregory.Argentieri-at-pharma.novartis.com } } Gregory.Argentieri-at-sandoz.com } } Greg2NJ-at-aol.com } } } } 201-503-8617Dear Greg, } The list you sent suggests that you may want to clarify your needs a } } little further before you go shopping. } 1. Do you want a fully integrated system or something which is more } modular? Quantimat and Kontron systems are sold as systems whereas } Media } Cy, Optimas, and Noeisis are software only. } 2. You mentioned the level of programming. I am more familiar with } the } off-the-shelf systems and of those, Noesis is the most powerful but } requires the greatest sophistication in terms of programming } ability. } Media Cy's Image Pro Plus and Optimas are more mid-range products. } We } have conducted a variety of market research projects, both at } meetings } (Microscopy & MicroAnalysis, ASMaterials, Experimental Bio, etc.), } by } phone, and personal interview. Of these two packages, Media Cy's } Image } Pro is the more widely accepted software. Several system } integrators } also indicated that it is more user friendly and the package has } been } adopted by a number of the microscope companies (ex: Zeiss' Image } One is } a privately labeled version; I expect that Nikon's offering is the } same, } Topcon and Phillips have used IPP in conjunction with their EMs). } 3. Re: the hardware - we strongly suggest that you talk to a few } local } system integrators. They can give you the latest on the } hardware/software/camera technologies and put together a system } which } meets your needs. (Input from other colleagues never hurts, } however). } } Hope this helps. If you need training when you get your system } together, } give us a call. } } Barbara Foster } Consortium President } Microscopy/Microscopy Education } } We have no financial interest in any of the products mentioned } above.
Greg - since you are upgrading you might also look at Amerinex's Aphelon software. It seems to be very powerful being a spinoff of some AI software developed for the military. I believe that they are at http://www.amerinex.com. If you are willing to program a system youmight also look at ContextVision's MicroGOP - they are from Sweden and the only number I have is there - 46-13-102480 or FAX 46-13-104282.
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4/22/97 4:25 PM Ultramicroscopy
Does anyone know what's become of the Elsevier journal "Ultramicroscopy?" They have apparently not published a single issue in 1997, although they normally appear monthly. An inquiry with the publisher was answered with a brief statement "Thank you for your inquiry. Unfortunately, the journal has not yet been published for 1997." I would withdraw my paper so that it could be published elsewhere (it was accepted in December 1996, and I even reviewed the galley proofs)... but it was supposed to be part of a conference proceedings issue (the Sixth Frontiers Conference, June 1996), so I would like to find out more before doing anything.
Still working in the dark... Jeff Fortner Argonne National Laboratory Chemical Technology Division Argonne, IL 60440
I am looking for a film scanner (digitizer for HRTEM films), which has resolution better than 2000 pixels/inch and 12 or more bits per pixel. I found one from Kodak's products, but it is too expensive (it works for color film, too). Anyone who has suggestion please email to panx-at-umich.edu Thank you.
Xiaoqing
_______________ Xiaoqing Pan, Ph.D. Associate Professor, Materials Science & Engineering University of Michigan 2038 H. H. Dow Building 2300 Hayward Street Ann Arbor, MI 48109-2136
Xiaoqing Pan wrote: } } I am looking for a film scanner (digitizer for HRTEM films), which } has resolution better than 2000 pixels/inch and 12 or more bits } per pixel. I found one from Kodak's products, but it is too expensive.
Dear Xiaoqing,
I have used an Agfa SelectScan, with specs (from my local distributor): Optical Resolution 4000 ppi x 4000 ppi Optical Density Range 3.6 Bits per pixel 13 (samples at 16) Cost ~A$72,000
I had no trouble with the digitised images from this scanner (other than the amount of RAM required to manipulate them!) ... a satisfied customer.
Given the high price of the SelectScan, another option might be the Agfa Vision 35: Optical Resolution 3175 ppi x 3175 ppi Optical Density Range 3.2 Bits per pixel 12 Cost ~A$12,000
I have no commercial interest in either of these products.
Stephen. ............................................................... : Stephen Anderson Australian Key Centre for : : Microscopy and Microanalysis : : Email stephen-at-emu.usyd.edu.au The University of Sydney : : Telephone (+61)-2-9351 7552 NSW 2006 : : Facsimile (+61)-2-9351 7682 Australia : :.............................................................:
We will hopefully get funding soon for a mid range voltage (300, 400 kV) TEM such as the H-9000, 3010, 4010 or CM300. We are very interested in obtaining private warts and all reports from operators of such microscopes on their satisfaction with their purchase.
We are interested in answers to questions such as:
What microscope did you choose and what was the main reason for your selection?
From the start of installation, how many days were taken before resolution was confirmed?
In days per year, how often is the microscope unusable with instrument failure?
Have there been any catastrophic ($10,000 +) failures?
Is it easy to maintain specified resolution?
How easy is it to train users to operate?
Would your microscope fit easily into a multi-user laboratory?
Are there any serious design problems you know of?
Would you buy another microscope from the same maker?
How would you rate your overall satisfaction with your microscope?
1 2 3 4 5 very unsatisfied unsatisfied just satisfied quite satisfied ecstatic
Mail replies to me, rather than posting them. I will collate replies and if anyone else wants to know the score, they can mail me for a private copy of the report.
Mel Dickson Director, E.M. Unit, University of New South Wales, Sydney Australia
At HRL we are urgently seeking a circuit board for the Motorised Stage Drive (ISM-MSD40-2) for our Jeol 840 SEM. The required circuit board is the computer interface (RS232) for external control, ie a EDXA control of the SEM stage. Should anyone know the whereabouts of a spare board or can get the schematic of this board we will be glad to here from you.
I have a colleague who would like to know if there is a way to deside if bacteria in activated sludge are gram-positive or gram-negative. Fresh samples were mixed with glycerol and frozen, thawed and prepared for SEM and TEM ( GA, OsO4, epoxy). There are no obvious signs of either gram-pos or neg. Is there any stain for sections? Fresh samples are no longer available.
TIA
Gunnel Karlsson
^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^ Gunnel Karlsson E-mail Gunnel.Karlsson-at-oorg2.lth.se Biomicroscopy Unit Tel +46 222 8229 Inorganic Chemistry 2 Fax +46 222 4012 Box 124 S-221 00 LUND, Sweden ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^ This message was sent by Eudora with recycled electrons
Hi Folks: We would like to do some standard processing and feature extraction from large images (about 10 megs, 8bit gray scale). Essentially we will use one image set to create a mask for feature extraction in another. The question is which packages are able to do this? I imagine the Photoshop image processing package (discussed a few weeks ago) will be able to generate the masks, however will this perform any quantitative extraction. Our current packages are limited by the frame size of the frame grabber, we would like to do this offline using standard ram memory. The platform can be PC, SGI or Mac.
Thanks
Simon C. Watkins Ph.D. Associate Professor and Director CBI University of Pittsburgh Pittsburgh PA 15261 tel:412-648-3051 fax:412-648-8330
Ultramicroscopy did exactly the same thing last year -- claiming "technical" problems then not appearing for many months. Yesterday I recieved my first/only copy of the year. I hope others will express (to elsevier) their frustration/anger at what used to be a good journal, but is now so unreliable as to be near useless. We are the customer, and the customer is right (or should be).
++++++++++++++++++++++++++++++++++++++++++++++++ Laurence Marks Department of Materials Science and Engineering Northwestern University Evanston, IL 60208-3108 tel: (847) 491-3996 fax: (847) 491-7820 email: l-marks-at-nwu.edu http: //www.numis.nwu.edu ++++++++++++++++++++++++++++++++++++++++++++++++
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Has anyone encountered oil contamination problems on cooled TEM ccd cameras? We recently fitted a Gatan MSC 791 to the 35mm port of a Hitachi H7100 TEM and have encountered massive oil contamination on the cooled ccd YAG. Several solutions have been suggested: fit foreline traps to the RP pumps to minimise backstreaming (done), fit a cold trap to the camera chamber diff. pump (may not be enough space), fit a turbo pump (cost issues), fit a cold trap inside the camera chamber (?) or modify the way the camera is run e.g. cooled in use, then immediately switched to warm cycle and leave at ambient when not in use. Any suggestions on the best way to resolve this problem would be much appreciated - also any general information on how others run cooled cameras would be useful - continuously cooled or only when in use, time intervals between heating cycles, camera temperatures, oils used etc........ Thanks in anticipation
Kevin Jennings
SmithKline Beecham Pharmaceuticals Harlow Essex U K
} 4:25 PM } Ultramicroscopy } } Does anyone know what's become of the Elsevier journal "Ultramicroscopy?" ...
snips ...
} Still working in the dark... } Jeff Fortner } Argonne National Laboratory } Chemical Technology Division } Argonne, IL 60440
I undertake a regular literature review, every 2 months. The last issue of Ultramicroscopy I saw was 65(3/4) October 96, and the last time I was in the library was early April.
I have found similar problems with a number of other journals in the microscopy area. Personally, I would suggest that if publishers can't supply journals on a regular basis, the journal becomes irrelevant. Surely, major objective of science and publication in journals is about communicating information - if a nominally monthly journal doesn't appear for 6 months, then perhaps, unless reasonable explanations are provided, it is time to withdraw papers, cancel subscriptions and request refunds?
} I am looking for a film scanner (digitizer for HRTEM films), which } has resolution better than 2000 pixels/inch and 12 or more bits } per pixel. I found one from Kodak's products, but it is too expensive } (it works for color film, too). Anyone who has suggestion please } email to panx-at-umich.edu } Thank you. } Dear Xiaoqing, If you are doing quantitative work, you should consider something like the Perkin-Elmer or Optronics. (There may be other spot-scanners, but I don't know what brands they are.) The P-E has a 5 micron square aperture setting, and the scanning densitometers are *much* more accurate--especially with negs having large dynamic range over small distances. Stray light landing on a particular pixel in a CCD array can be significant if the pixel you're measuring is very dark, and this problem increases the nearer the bright pixels are to the dark ones. The low-order spots in an ED pat- tern, for example, have OD's of up to ~4, and proper background subtraction can only be performed if these are measured accurately. For quantitative comparisons of images to simulations, you may need similar accuracy, and, although you might be able to fold a "scanner contrast transfer function" into your simulation, it is better to get the measurements right in the first place. We have a P-E, but I have no financial interest in any of this. Yours, Bill Tivol
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Reply to: RE} Ultramicroscopy
We have taken contact with the editor in chief about the 97 volumes of Ultramicroscopy , Pieter Kruit , and the situation is as follows: After changing to computer aided production the publisher , Elseviers had a delay of several months Due to this the normally planned five volumes of 96 were not all published in that year. In order to keep their obligations the last volumes of 96 will come out in may 97 (They all contain material that was submitted in 96) From the end of may the new 97 volumes will appear ; Dirk van dyck
A Gatan 694 camera attached to the bottom of the camera chamber of our dry-pumped Hitachi HF-2000 initially exhibited quite bad oil contamination. After a couple of times removing the camera and cleaning the surface of the detector with a gentle wash of freon, the problem basically disappeared. We are convinced that the problem was caused by wicking of the oil film between the two fiber optic plates. Perhaps there was an initial excess of oil used to couple the plates together. Since there is no significant source of oil contamination in our microscope, other than what might come from O-rings, this seemed to be the only explanation. Interestingly, a similar camera on our JEOL 4000EX, which has a DP on the camera chamber, has not had any contamination problems. I think that if you have backstreaming sufficient to cause the problem on the CCD, you will have other contamination problems also....
Larry
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Why such large images (3000x3000 pixels)? Just getting and storing the images would seem a tremendous challenge. We routinely work with 1 MB images in Visilog. At that large stage, I would wonder about doing a hybrid system where the whole image is not stored. Detect the particles in a live mode and analyze them live or store just the area for your features of interest. Of course, this only works well with random access to all parts of the field.
I began image analysis with a 64 KB computer running hardware and software from LeMont Scientific. All intensities were read live from our SEM. Only the measurements were stored. Later versions also worked off of stored images. I have also seen a package from the R.J.Lee Group (Monroeville, PA, your back yard) which took a somewhat similar approach. Much of the measurement was done live, but images were also stored for each feature.
Disclaimer, I have no financial interest in the above companies. I remain fascinated by their ingenuity.
At 07:54 AM 4/23/97 -0400, Simon Watkins wrote: } ------------------------------------------------------------------------ } } Hi Folks: } We would like to do some standard processing and feature extraction from } large images (about 10 megs, 8bit gray scale). Essentially we will use one } image set to create a mask for feature extraction in another. The question } is which packages are able to do this? I imagine the Photoshop image } processing package (discussed a few weeks ago) will be able to generate the } masks, however will this perform any quantitative extraction. Our current } packages are limited by the frame size of the frame grabber, we would like } to do this offline using standard ram memory. The platform can be PC, SGI } or Mac. } } Thanks } } } Simon C. Watkins Ph.D. } Associate Professor and Director CBI } University of Pittsburgh } Pittsburgh PA 15261 } tel:412-648-3051 } fax:412-648-8330 ---------------------------------------------------- Warren E. Straszheim 270 Metals Development, Ames Lab/ISU, Ames IA, 50011 Phone: 515-294-8187 FAX: 515-294-3091
} I have a colleague who would like to know if there is a way to deside if } bacteria in activated sludge are gram-positive or gram-negative. Fresh } samples were mixed with glycerol and frozen, thawed and prepared for SEM } and TEM ( GA, OsO4, epoxy). There are no obvious signs of either gram-pos } or neg. Is there any stain for sections?
Gram negative can readily be distinguished from gram positive bacteria by examination of the structure of the cell wall by TEM. Check any good general bacteriology book for figures. The outermost layers ("wall") of gram negs show what appear to be two unit membranes with a thin (usually dense) amorphous layer between. This outermost membrane (containing lipopolysaccharide -} common in G- but extremely rare in G+) is most often undulated, giving the appearance of a ruffled surface by SEM. Gram positives, on the other hand, have a single cell membrane with a thick (usually electron-light) wall exterior to the membrane. The wall is composed of peptidoglycan and lacks LPS or lipopolysaccharide. If you need more info or require a specific reference, contact me.
#################################################################### John J. Bozzola, Ph.D., Director Center for Electron Microscopy Neckers Building, Room 146 - B Wing Southern Illinois University Carbondale, IL 62901-4402 U.S.A. Phone: 618-453-3730 Fax: 618-453-2665 Email: bozzola-at-siu.edu Web: http://www.siu.edu/departments/shops/cem.html ####################################################################
} Has anyone encountered oil contamination problems on cooled TEM ccd } cameras? We } recently fitted a Gatan MSC 791 to the 35mm port of a Hitachi H7100 TEM and } have encountered massive oil contamination on the cooled ccd YAG. Several } solutions have been suggested: fit foreline traps to the RP pumps to minimise } backstreaming (done), fit a cold trap to the camera chamber diff. pump } (may not } be enough space), fit a turbo pump (cost issues), fit a cold trap inside the } camera chamber (?) or modify the way the camera is run e.g. cooled in use, } then } immediately switched to warm cycle and leave at ambient when not in use. Any } suggestions on the best way to resolve this problem would be much } appreciated - } also any general information on how others run cooled cameras would be } useful - } continuously cooled or only when in use, time intervals between heating } cycles, } camera temperatures, oils used etc........
Sounds like you have a vacuum problem with the EM. You should NOT be seeing "massive" oil contamination in this generation of microscope. Call the EM service people because the oil is going elsewhere in the EM as well.
#################################################################### John J. Bozzola, Ph.D., Director Center for Electron Microscopy Neckers Building, Room 146 - B Wing Southern Illinois University Carbondale, IL 62901-4402 U.S.A. Phone: 618-453-3730 Fax: 618-453-2665 Email: bozzola-at-siu.edu Web: http://www.siu.edu/departments/shops/cem.html ####################################################################
Greetings collective, and special thanks to Nestor for clearing up those pesky double posts..
I'm working up an experiment that will compare freeze drying and freeze substitution on some Geranium pedicel trichomes. I would like to embedd in GMA and photo polymerize at a cool temperature, probably ca. 5C. After sectioning, I will de-embed and probe the sections with our imaging mass spect instrument (TOF-SIMS) to see how the chemical constituents have fared.
This is my first attempt with a GMA kit (EMS) and cold embedding/polymerizattion. Any tips from seasoned users.....? TIA
Edward J. Basgall, PhD The Pennsylvania State University Surface Chemistry Group ejb11-at-psu.edu Materials Research Institute Building Ph: 814-865-0493 University Park, PA 16802-7003 FAX: 814-863-0618
Up to the age of forty, eating is beneficial; after forty, drinking. - The Talmud
TO WHOM IT MAY CONCERN, I heard that I could get some answers to my preperation problems here. I have been trying to prepare sickle cells for the SEM and I keep getting artifacts like echinocytes and folded cells. I would like to know if anyone could send me some special preperation instructions that I could use. Any help at all would be greatly appreciated. Thank you Sincerely, Emmanueuel Uche
I would like to announce an opening for - Senior Analyst/Microscipist; as of July 1, 1997.
An immediate position is open for a senior analyst at the North Carolina State University Analytical Instrumentation Facility (AIF) as a retirement replacement.
Duties and responsibilities include: operation and maintenance of optical metallographs, ion millers, X-Ray diffractometers and sample preparation devices such as mounting presses and grinding, polishing and sectioning devices, etc; scheduling of access to and oversight of the above instrumentation; and user training and assistance. Other responsibilities include operation of SEM and TEM and assistance with the teaching of electron microscopy laboratory classes and assistance with other graduate level engineering classes. Qualifications must include an BS as the minimum degree with higher degree desired or equivalent experience in a materials related discipline (non biological) along with hands on analytical experience in a multiuser analytical facility environment,
Please send resume and names of references to: Phil Russell, Director; Analytical Instrumentation Facility; North Carolina State University; Box 7531, Room 118A EGRC; 1020 Main Campus Drive; Raleigh, NC 27695-7531 (e-mail is acceptable; to prussell-at-ncsu.edu)
North Carolina State University is an Equal Opportunity, Affirmative Action Educator and Employer. Minority and Female Applicants are especially encouraged.
Phillip E. Russell Analytical Instrumentation Facility Box 7531, Room 318 EGRC North Carolina State University Raleigh, NC 27695-7531
A thread similar to this has been archive at the "Tips & Tricks " site. Go to the web address at the end of this message and look for the Tips & Tricks link. Go to the TEM section or use the search utility and you will find the replies to the earlier posting. Good luck
At 10:05 AM 4/23/97 +0200, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { GO GATORS Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 352-392-1295 ICBR EM Core Lab fax 352-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/ The home of " Tips & Tricks "
OOPS, sorry folks. The kit I will use is a methyl methacrylate/butyl methacrylate kit not GMA as I posted earlier.....
} I'm working up an experiment that will compare freeze drying and freeze } substitution on some } Geranium pedicel trichomes. I would like to embed } in MBM and photo polymerize at a cool } temperature, probably ca. 5C. } After sectioning, I will de-embed and probe the sections with our } imaging } mass spect instrument (TOF-SIMS) to see how the chemical constituents have } fared.
} This is my first attempt with a MBM kit (EMS) and cold } embedding/polymerizattion. Any tips } from seasoned users.....? } TIA
Edward J. Basgall, PhD The Pennsylvania State University Surface Chemistry Group ejb11-at-psu.edu Materials Research Institute Building Ph: 814-865-0493 University Park, PA 16802-7003 FAX: 814-863-0618
If you really want a powerful, not necessary user friendly image processing package. I suggest you try semper6. Basically it has no limits of the image size. I use it to analysis images in the lab. Feel it is the one I most liked. Semper6 is a script language package. Not for dummy but for scientists. It has SGI, SUN and PC's versions. The problem is, it is not cheap. But you can get a demo version to try with.
I have no association with Semper6. It is developed by Cambridge U. (?)of UK.
I'm working up an experiment that will compare freeze drying and freeze substitution on some Geranium pedicel trichomes. I would like to embed in MBM and photo polymerize at a cool temperature, probably ca. 5C. After sectioning, I will de-embed and probe the sections with our imaging mass spect instrument (TOF-SIMS) to see how the chemical constituents have fared.
This is my first attempt with a MBM kit (EMS) and cold embedding/polymerizattion. Any tips from seasoned users.....? TIA
Edward J. Basgall, PhD The Pennsylvania State University Surface Chemistry Group ejb11-at-psu.edu Materials Research Institute Building Ph: 814-865-0493 University Park, PA 16802-7003 FAX: 814-863-0618
We have a new Kevex Sigma 3 system with the Mirage image analysis software. Is there anyone out in EM land with a step-by-step protocol/instructions for imaging and analysis on Mirage? We are having difficulty deciphering the manual. Replies can be emailed to me or better yet, the instructions FAXed to me at 515.294.1337. My phone number is 515.294.3872. Thank you in advance!! Bruce Wagner.
We have the same system you are asking about and I had similar problems. If you think the Mirage manual is bad, you should have seen the manual for the Visilog 4 system I started with. The best advice I can give you is to read John Russ's "The Image Processing Handbook" or some similar text. If there is anything specific I can help you with, give me a call or send me an email.
John Robinson Electron Optics Technologist Mechanical & Materials Engineering University of Windsor Windsor, Ontario N9B 3P4
Hello, Lookng for the voice(s) of experienced TEM users (as opposed to vendors). What is a reasonable expectation for stage stability in a late model 200KV TEM. say in nm/min.?
I am interested in any hint in order to perform the calibration of a TEM high temperature sample holder test, up to 1000 degrees C. The idea is looking at various sample with a known structural transformation ocurring at precise temperature, and compare this "real" temperature with the one given by the sample holder thermocouple. Does anyboby have performed such a test before, and with which materials. I believe that having 4 or 5 points between 100 and 1000 degrees would be a good start.
thanks,
Yves MANIETTE Universitat de Barcelona Serveis Cientifico Tecnics Unitat ESCA TEM Carrer Lluis Sole i Sabaris, 1-3 E-08028 BARCELONA ESPANYA
Unfortunately, calibrating the electron-transparent portion of the TEM specimen to the TEM specimen holder temperature will be valid only for that particular specimen loaded at that particular time. Take the specimen out, and the former calibration will loose accuracy. This is due to the fact that you will not be able to duplicate the way in which you load (clamp) the specimen, so that the thermal contact resistance (between the specimen and specimen cup) will change. Fortunately, this likely amounts to only a few degrees of error. Furthermore, any calibration should only be used for similar specimens of similar thickness and having similar specimen preparation. If you calibrate the TEM holder using a specimen of relatively poor thermal conductivity, don't expect it to be correct for a specimen having high thermal conductivity. All bets are off if the specimen is not homogeneous in thermal conductivity (i.e., if the specimen has a crack in it, or is a multilayer, as the resistance to heat flow can vary locally so that the actual temperature can be significantly below the furnace temperature).
Its an interesting problem. We used the regrowth rate of amorphous silicon to establish the local specimen temperature in Si-Ge in:
D. C. Paine, D. J. Howard, N. D. Evans, D. W. Greve, M. Racanelli, and N. G. Stoffel, "In Situ TEM Studies of the Growth of Strained Si1-xGex by Solid Phase Epitaxy," in Evolution of Thin Film and Surface Microstructure, Mater. Res. Soc. Symp. Proc. 202, C. V. Thompson, J. Y. Tsao, and D. J. Srolovitz, eds., Materials Research Society, Pittsburgh, PA (1990)
} Date: Wed, 23 Apr 1997 12:49:15 -0700 (PDT) } From: Emmanuel Uche {euche-at-haywire.csuhayward.edu} } To: Microscopy Headquarters {microscopy-at-sparc5.microscopy.com} } Subject: NEED HELP W/ BLOOD SAMPLES } } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } TO WHOM IT MAY CONCERN, } I heard that I could get some answers to my preperation problems } here. I have been trying to prepare sickle cells for the SEM and I keep } getting artifacts like echinocytes and folded cells. I would like to know } if anyone could send me some special preperation instructions that I could } use. Any help at all would be greatly appreciated. Thank you } Sincerely, } Emmanueuel Uche } A long time ago, we did some work on RBC's from muscular dystrophy patients. We found that the cells are very sensitive to pH, salt concentration, glass, and other things, maybe including the moon??? Low pH, increased salt, esp. NaCl, and sitting in a glass tube or on a glass slide unfixed would cause echinocytes. We got around the problem by drawing the blood and then IMMEDIATELY dropping (drop by drop) blood samples into 1% glutaraldehyde at pH 7.4, in 300 mOSM phosphate buffer, not PBS. Test the osmolarity of the buffer; the final osmolarity of the buffered fix will be 400: 300 from the buffer and 100 from the glut! The fix should be room temperature (not cold). Only about 2-3 drops of blood should be put into 10 ml fix. Gently invert several times to mix, but don't shake vigorously. Let them sit in the tube, and they will settle out to the bottom, or you can put a drop of suspension onto a collagen-coated coverslip, allow them to settle out for an hour (cover in a moist champer), then gently dehydrate in ethanol, and critical point dry. I don't remember which references contain the method, but try these:
Miller, S. E., A. D. Roses and S. H. Appel. 1975. Erythrocytes in human muscular dystrophy. Science 188:1131.
Roses, A. D., S. H. Appel, D. A. Butterfield, S. E. Miller and D. B. Chesnut. 1975. Specificity of biochemical and biophysical tests in Duchenne and myotonic muscular dystrophy, carrier states and congenital myotonia. Trans. Amer. Neurol. Assoc. 100:131-134.
Roses, A. D., M. J. Roses, S. E. Miller, K. J. Hull, Jr. and S. H. Appel. 1976. Carrier detection in Duchenne muscular dystrophy. New Engl. J. Med. 294:193-198.
Miller, S. E., A. D. Roses and S. H. Appel 1976. Scanning electron microscopy studies in muscular dystrophy. Arch. Neurol. 33:172-174.
Good luck, SM
Sara E. Miller, Ph. D. P. O. Box 3020 Duke University Medical Center Durham, NC 27710 Ph: 919 684-3452 FAX: 919 684-8735
Dear list, We are looking for slides imprinted with grid squares so we can maintain orientation/coordinates. I was wondering if anyone has knowledge of any source that carries slides marked for orientation. Any help will be appreciated. with regards,
************************* * Marilyn Wadsworth * * mwadswor-at-zoo.uvm.edu * *************************
I am still having problems with my cpd efforts giving me 'raisins' especially on young plant tissue. Has anyone used HMDS? How do the results compare to CPD, or Freon (I know I can't use freon, but it used to be the standard)?
Are there any tricks I should know before embarking on a HMDS experiment?
Thanks for letting me pick your brains.
Paula = )
Paula Sicurello UC Berkeley Electron Microscope Lab psic-at-uclink4.berkeley.edu
} Lookng for the voice(s) of experienced TEM users (as opposed to vendors). } What is a reasonable expectation for stage stability in a late model 200KV } TEM. say in nm/min.? } Dear Bruce, Not quite the answer to your question, but... The HVEM has a long- term drift of a few nm/min--I didn't have the time to look up our test re- sults. We test by inserting a stage, letting everything sit for ~1/2 hour and checking at 15 min intervals to see how far the image has moved. We found remarkably little movement until we looked more closely. It turned out that our Haskris chillers cycled with a period close to the observation interval, and when we looked continuously, we saw a cyclic drift pattern. We ordered a hot-gas-bypass unit for the chillers, and this gave much more stability. If your stability results are } } a few nm/min, you might look at some of your ancillary equipment (including room heating & air condi- tioning, etc.) to see if there is anything outside the scope causing in- stabilities. Good luck. Yours, Bill Tivol
This is a REPOSTING of the OFFICIAL PROPOSAL for a NEW USENET NEWSGROUP that will probably be named "sci.bio.immunocytochem" formally posted to news.announce.newgroups on 16.4.97.
Immunocytochemists/immunohistochemists may like to read the following proposal and post their comments or suggestions to "news.groups" where the set-up discussion is taking place, or you may e-mail me with your comments and I will repost them for you {awilson-at-aw.u-net.com}
REQUEST FOR DISCUSSION (RFD) unmoderated group sci.bio.immunocytochem
This is the 3rd Request For Discussion (RFD) for the creation of a world-wide unmoderated Usenet newsgroup sci.bio.immunocytochem, currently being discussed in news.groups
Suggestions for improvements to this proposal are welcome. Discussion about it should take place in news.groups. A vote is expected to be held in about three weeks.
This is not a Call for Votes (CFV); you cannot vote at this time. Procedural details are below.
CHANGES from previous RFD:
There is an important change to the Charter of this newsgroup: It is proposed that the new group will include the discussion of related affinity labelling methods as well as immunohistochemical and immunocytochemical topics. Minor changes to rational too.
Newsgroup line: sci.bio.immunocytochem Immuno-labelling of biological material.
RATIONALE: sci.bio.immunocytochem
Immunohistochemists and immunocytochemists already enjoy the benefits of online communication, utilizing e-mail, accessing web sites, and subscribing to specialised mailing lists. Usenet newsgroups are also popular, but this is less obvious because articles with immunocytochemical/immunohistochemical content get posted to many different newsgroups. Most articles are posted to a favourite five or six newsgroups including bionet.cellbiol, sci.med.immunology and sci.techniques.microscopy, but often articles get posted to any one of fourteen or fifteen newsgroups in the sci. and bionet. heirarchies. Some of these are listed in the distribution list at the end of this proposal.
In my view, no existing newsgroup fulfils the criteria necessary to attract all the various immunocytochemistry postings. I do not wish to draw users away from other newsgroups, only to encourage scientists to share their knowledge and expertise on immunocytochemistry in the most effective manner.
Immunocytochemistry and immunohistochemistry are not subdivisions of immunology, molecular biology or chemistry. Microscopy, although essential, is only a small part of the story. Immunocytochemistry and immunohistochemistry are multi- disciplinary, therefore discussions are destined to stay distributed amongst the different newsgroups until they are all brought together under one umbrella. This would then act as a focus point for all the immunocytochemists who are already Internet users, and encourage new subscribers to Usenet.
CHARTER: sci.bio.immunocytochem
This is a newsgroup for the exchange of information relating to immunocytochemistry and immunohistochemistry, and all forms of related affinity labelling methods, such as lectins and in-situ hybridisation. These unique research tools are used to locate and identify specific molecules in biological material, at the microscopical level.
Articles posted to this group must be relevant to one or more aspects of the above. The kind of subjects that may be discussed include techniques, theory, presentation of results, requests for collaboration, history, equipment, publication references, notice of events, tips and trouble-shooting, jobs offered andwanted, jokes, stories and new ideas, so long as the posting bears a direct relevance to the central theme. There will be a list of Frequently Asked Questions (FAQs) to help newcomers.
A relevant posting could just be a simple question or answer, for example "Has anyone got any experience with this reagent ?"or "Which course could I attend to learn more about immunogold labelling?". There will be articles reminding people to read the list of FAQs prior to posting their own article. Usenet readers may get involved in complex discussions about, for example, multiple labelling, proper use of control experiments, microwave antigen retrieval or quantitative measurements. Remember that articles posted to a newsgroup are intended for a wide readership, so if you have information which concerns only one or two people then please don't use this newsgroup, use e-mail.
Commercial advertisements for services, equipment or reagents violate the charter unless one or more of the following apply: (a)The advertisement is part of a comprehensive article designed specifically to address issues raised in earlier articles posted to the group (b)A general reference to the type of product does not suffice for technical reasons and it is necessary to specify the exact commercial product (c) The information is offered primarily for the benefit of the readers (d)The advertisement is for second-hand equipment specific to immunocytochemistry (e) Requests or offers for free products are acceptable if they are not part of a sales promotion.
END CHARTER.
PROCEDURE:
This is a request for discussion, not a call for votes. In this phase of the process, any potential problems with the proposed newsgroups should be raised and resolved. The discussion period will continue for a minimum of 21 days (starting from when the 3rd RFD for this proposal is posted to news.announce.newgroups), after which a Call For Votes (CFV) may be posted by a neutral vote taker if the discussion warrants it. Please do not attempt to vote until this happens.
All discussion of this proposal should be posted to news.groups. This RFD attempts to comply fully with the Usenet newsgroup creation guidelines outlined in "How to Create a New Usenet Newsgroup" and "How to Format and Submit a New Group Proposal". Please refer to these documents (available in news.announce.newgroups) if you have any questions about the process.
DISTRIBUTION:
This RFD has been posted to the following newsgroups: news.announce.newgroups,news.groups,bionet.cellbiol, bionet.diagnostics,bionet.immunology,bionet.microbiology, bionet.molbio.methds-reagnts,sci.bio.misc,sci.med.immunology, sci.techniques.microscopy
This RFD will be reposted to the following newsgroups after its posting in news.announce.newgroups: bionet.molbio.proteins,bionet.neuroscience,bionet.plants, sci.bio.microbiology,sci.med,sci.med.laboratory,sci.misc, sci.nanotech
This RFD will also be reposted to the following mailing lists after its posting in news.announce.newgroups:
Histonet mailing list: {histonet-at-pathology.swmed.edu} Information pertaining to the technical aspects of histology and histopathology such as tissue fixatives and processing, routine histology, special stains, immunohistochemistry, in-situ hybridization etc. To subscribe type "subscribe digest" into the subject box and leave the text box empty, or to subscribe to the full service just type "subscribe". For more info access web site http://www.mwrn.com/subject/histonet.htms
Microscopy Society of America listserver: Questions/comments/answers in the various fields of Microscopy Currently over 3000 subscribers. To subscribe send the message "subscribe" to {Listserver-at-MSA.Microscopy.Com} then send messages in plain text to {Microscopy-at-MSA.Microscopy.Com} For more info access web site http://www.amc.anl.gov/ Docs/anl/Nestor/Software/telecommList.html
Stanford University list server To subscribe, send a message to {majordomo-at-pathology.stanford.edu} with "subscribe ipox-l" in the body of your message. This list helps pathologists and other laboratory professionals to exchange information about immunoperoxidase methods.
This RFD will also be reposted to the following web-sites after its posting in news.announce.newgroups:
Royal Microscope Society http://www.rms.org.uk Web Master Dr R. A. D. Mackenzie {r.a.mackenzie-at-open.ac.uk}
Center for Cell Imaging Department of Cell Biology Yale University School of Medicine Introduction to Immunocytochemistry http://info.med.yale.edu/cellimg/CCIimmuno.html Web Master Paul Webster { paul_webster-at-yale.edu}
Proponent: Amanda Wilson {awilson-at-aw.u-net.com} Proponent: Paul Monaghan { monaghan-at-icr.ac.uk} Mentor: Jonathan Grobe {grobe-at-netins.net}
To all those folks who have a violent interest in Pb stains for TEM, and to all those who have requested a copy of the book chapter -
I am sorry to have have had to delay my answers to you all on this most intriguing topic - we have been suffering here from MEGA grant insanity. On Tuesday, April 29, I will plan to post a message and bundle up the book chapter copies. Thanks for your patience. Hildy
Hi, Although I guess I could be considered a vendor; I have been known to use a microscope from time to time... The question you ask is not complete; I could reply that any microscope stage which does not use sliding 'O' rings can be seen to have drift rates well below 1 nm/min, depending on how much you are prepared to spend on the site. It is very much a factor of the temperature stability of the sample in the holder, the holder/sample temperature, and the sample temperature when inserted should be as equal to the in-column temperature as possible to give the best results. When you have all the thermal factors pretty well sorted out (now you've spent the best part of $100,000 on THE ROOM!!) i.e. no drafts and no difference in temperature between the sample whether it is inside the column or outside the column, THEN (and only then) can you start to think about what happens when you turn on the beam. IF your sample conducts heat reasonably, then you have a chance at good stability below 3 angstroms/min. There are not many vendors who would endorse such numbers because of the site variables that are involved. If the sample has poor conductivity, then the beam will cause local heating and then the sample will move no matter what you try to do.
Any other inputs on this subject would be much appreciated. I know that Tim Baker at Purdue has measured some unbelievable drift rates using a cold stage (COLD!) but I cannot quote them here...
Cheers, Jan
Dr. Jan Ringnalda Sr. Apllications Specialist, FEI/Philips Electron Optics, 85 McKee Dr., Mahwah, NJ 07430 (201) 529 6160
Dear Bruce, From my own experience, the sample should drift several nm /min for the first 1/2 hour, then settle down to little or no drift after that, unless the specimen itself moves under the beam. Speaking of voices, if you observe the image at ~200,000 and talk to someone, you will see the specimen vibrate when your voice hits certain frequencies. Remember not to talk while you are photographing. A microscope designer told me they once improved stage stability by reducing the diameter of the handle of the specimen holder, so it didn't react to air drafts and thermal gradient so much. This is why high resolution TEMs are top entry. Air drafts, air conditioners, noisy water chillers, any other noise or source of drafts or thermal changes will affect the stage. You wrote:
} Hello, } Lookng for the voice(s) of experienced TEM users (as opposed to vendors). } What is a reasonable expectation for stage stability in a late model 200KV } TEM. say in nm/min.?
Hope this helps, Mary Mary Mager Electron Microscopist Metals and Materials Eng., UBC 6350 Stores Rd. Vancouver, B.C. V6T 1Z4 CANADA tel:604-822-5648, fax:604-822-3619 e-mail: mager-at-unixg.ubc.ca
} We would like to do some standard processing and feature extraction from } large images (about 10 megs, 8bit gray scale). Essentially we will use one } image set to create a mask for feature extraction in another. The question } is which packages are able to do this? I imagine the Photoshop image } processing package (discussed a few weeks ago) will be able to generate the } masks, however will this perform any quantitative extraction. Our current } packages are limited by the frame size of the frame grabber, we would like } to do this offline using standard ram memory. The platform can be PC, SGI } or Mac.
I use Khoros2, which comes as a shareware 'Advanced' version and a commercial 'Pro' version (I think the differences are minor, but I only know the 'Advanced'). It runs on any computer running some sort of Unix (Sun, SGI, PC ...). I've processed byte images up to 4096 pixels on a Sun with 96 MByte RAM. Khoros will do almost anything You can imagine (or at least I can) in the field of general image processing. Check: http://www.khoral.com/
Hope that helps, -- Philip Koeck Karolinska Institutet Dept. of Bioscience Novum S-14157 Huddinge Sweden Tel.: +46-8-608 91 86 Fax.: +46-8-608 92 90 Email: Philip.Koeck-at-csb.ki.se http://www_scem.csb.ki.se/pages/philip.html
} I think Larry might be right in his case but that John B. is probably right } in your case. There is no way to tell, though, without knowing more about } the extent and duration or your problem. Could you elaborate on your } problem and include camera serial number and date shipped (if known)?
} Paul
Thanks to everyone for the comments so far. Here are more details - First noticed the problem after a couple of weeks use - overnight heating appeared to remove the contamination but it continued to reappear in the same position by the end of the day with continuous cooling. The camera (serial number K6070403 delivered Jan 97) was removed by an engineer after one of the heating cycles and found to be contaminated with oil in the same position as seen on captured images. After cleaning, the camera was reassembled and all appeared OK - although on switching on the following morning the camera failed completely with no image. On checking the camera, more oil was found plus the whole ccd /Peltier assembly appeared loose. This should be fixed next week after which we will review the problem again. As for microscope oil problems - the H7100 appears to have low contamination rates in normal use and has given no obvious problems whilst we've been using film.
Kevin Jennings
SmithKline Beecham Pharmaceuticals Harlow Essex CM19 5AW U K
} I am interested in any hint in order to perform the calibration of a TEM } high temperature sample holder test, up to 1000 degrees C. The idea is } looking at various sample with a known structural transformation ocurring } at precise temperature, and compare this "real" temperature with the one } given by the sample holder thermocouple. Does anyboby have performed such } a test before, and with which materials. I believe that having 4 or 5 } points between 100 and 1000 degrees would be a good start. } } thanks, } } Yves MANIETTE } Universitat de Barcelona
While this sounds like a good approach, I'm not sure it would give you a better accuracy than you already have with the built-in thermocouple. I think a lot depends on how accurately you really do need to know the temperature? You might also have difficulties finding a suitable material (that is, you can make a TEM specimen) at the lower end of the temperature range.
Firstly, the temperature given for such transformations probably applies to bulk material. It is possible that in very thin TEM specimens, the transformation temperature may be different?
Secondly, in most cases, such transformations aren't instantaneous - they move through a material from nucleation sites. As a consequence, I would expect the transformation to occur over a small temperature range, rather than at a specific temperature. Of course, if the range is small, say 1 degree, this may be sufficient for your purposes.
Thirdly, the thermocouple is measuring the temperature not of the specimen, but of some point near to the specimen, either in the holder tip, or, more likely, in the specimen cup. Thermal conductivity and specimen clamping considerations mean that there will always be a thermal lag between the specimen and the thermocouple; there may also be a temperature gradient, although I guess that in most cases this would disappear given a suitable period of time. This effect may also be influenced by beam heating, and will be different for every specimen.
An alternative approach (which I have not personally tried, so I don't know if it would really work) may be to measure thermal expansion. Although electron diffraction is not too good at making an absolute measurement of lattice parameter, convergent beam methods, looking at HOLZ lines can in principle measure lattice parameter changes to 1 in 1000 or 10000. If you start with a material with precisely known room temperature lattice parameter and thermal expansion coefficients, you should be able to measure the lattice parameter at several temperatures and then calculate back to a value for the actual temperature. This approach has the advantage that you really are measuring the temperature of the specimen plus you can decide at which temperature points to calibrate, and how many. It might also allow you to do such a calibration in situ - that is, with the material in which you are really interested.
I would suggest you contact both Gatan and Oxford Instruments, who make commercial TEM heating holders. I don't know about Oxford, but at Gatan contact Reza Alani {ralani-at-gatan.com} , their Applications Manager.
It's not really a drift problem, but I recall several years ago that I used a Philips 301(ironically) with eucentric goniometer and the stage seemed to have a resonant frequency somewhere near human voice levels - at least that's what we thought. This meant that the best pictures were taken by the quietest people. I don't know if similar problems still arise but they could obviously make normal specimen drift difficult to standardise.
Malcolm Haswell University of Sunderland UK ----------
Hi, Although I guess I could be considered a vendor; I have been known to use a microscope from time to time... The question you ask is not complete; I could reply that any microscope stage which does not use sliding 'O' rings can be seen to have drift rates well below 1 nm/min, depending on how much you are prepared to spend on the site. It is very much a factor of the temperature stability of the sample in the holder, the holder/sample temperature, and the sample temperature when inserted should be as equal to the in-column temperature as possible to give the best results. When you have all the thermal factors pretty well sorted out (now you've spent the best part of $100,000 on THE ROOM!!) i.e. no drafts and no difference in temperature between the sample whether it is inside the column or outside the column, THEN (and only then) can you start to think about what happens when you turn on the beam. IF your sample conducts heat reasonably, then you have a chance at good stability below 3 angstroms/min. There are not many vendors who would endorse such numbers because of the site variables that are involved. If the sample has poor conductivity, then the beam will cause local heating and then the sample will move no matter what you try to do.
Any other inputs on this subject would be much appreciated. I know that Tim Baker at Purdue has measured some unbelievable drift rates using a cold stage (COLD!) but I cannot quote them here...
Cheers, Jan
Dr. Jan Ringnalda Sr. Apllications Specialist, FEI/Philips Electron Optics, 85 McKee Dr., Mahwah, NJ 07430 (201) 529 6160
I am looking for a secondary detector tube for JEOL JSM 35 in a good condition. Willing to pay a fair price for it. Please reply at my email address. Many many thanks for your anticipated co-operation.
Jitu Shah
Dr.Jitu Shah H.H. Wills Physics Laboratory, University of Bristol, Royal Fort, Tyndall Avenue, Bristol BS8 1TL. UK email: jss-at-siva.bristol.ac.uk Tel: 44 117 9288719 Fax: 44 117 9255624
Generally, HMDS does not work on plant tissue. During the SEM course we teach, we have made the students use both procedures ( we are such slave drivers) and there seems to be no way to tell which plants it will work on and which it won't. Have you tried long infiltrations??? Atr the end of the CPD run do you allow the gas bleed off slowly?? It usually takes us 20-30 min. to come to atm.
I maintain an archive of the biologic related threads posted to this list. You may find it at the web address listed at the end of this message. Go to the "Tips & Tricks" link and look in the "SEM" section. There are some threads you may find enlightening. Good luck!!
At 01:45 PM 4/24/97 -0700, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { GO GATORS Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 352-392-1295 ICBR EM Core Lab fax 352-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/ The home of " Tips & Tricks "
We have never had good luck with HMDS on plant material. We always CPD. Be sure that ou have gotten ridof all the water and solvent before drying Sniff the chamber when you open and see if you still smell solvent. If yes you need longer or more changes during CPD } } Dear smart people, } } } I am still having problems with my cpd efforts giving me 'raisins' } especially on young plant tissue. Has anyone used HMDS? How do the } results compare to CPD, or Freon (I know I can't use freon, but it used to } be the standard)? } } Are there any tricks I should know before embarking on a HMDS experiment? } } Thanks for letting me pick your brains. } } } Paula = ) } } Paula Sicurello } UC Berkeley } Electron Microscope Lab } psic-at-uclink4.berkeley.edu } } } } ******************************************************* G.W. Erdos, Ph.D. Phone: 352-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax: 352-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 http://www.biotech.ufl.edu/~emcl/ Home of the #1 Gators ***** "Many shall run to and fro, and knowledge shall be increased" Daniel 12:4
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Marilyn Wadsworth writes:
"We are looking for slides imprinted with grid squares so we can maintain orientation/coordinates. I was wondering if anyone has knowledge of any source that carries slides marked for orientation. Any help will be appreciated."
Dear Marilyn,
I do not know of glass slides that are etched but what you may be looking for are coverslips with etched grids.
You may wish to contact "Eppendorf" to ask about their "CELLocate" system. You get small glass, sterile coverslips, etched with a numbered grid. Paper maps are supplied to mark interesting locations.
Cells grow well on the substrate and the etched grid will transfer easily to epoxy resin after embedding.
Regards,
Paul Webster Center for Cell Imaging Yale School of Medicine http://info.med.yale.edu/cellimg
The Electron Microscopy Laboratory at New Mexico State University has an open position for an Electron Microscopy Specialist. The laboratory provides transmission, scanning and light microscopy services for the university research community. Qualifications: B.Sc. degree minimum, M.Sc. degree desirable, with at least 4 years of electron microscopy experience. The preferred candidate will have experience with scanning electron microscopy and x-ray analysis. Experience with digial image analysis is also desirable as is experience with flourescense microscopy and immunocytochemistry. The candidate must be competent with sample preparation techniques including vacuum evaporation, sputter coating, critical point drying, support film production, low temperature embedding, and photographic film processing and printing. He or she must be able to work well with research faculty, staff and students, and be able to train and instruct graduate and undergraduate students. Duties: operation and routine maintenance of transmission and scanning electron microscopes and associated equipment, fixation, embedding, ultra-thin sectioning, staining, coating of samples, and record keeping. Salary: $26,872 to $30,972 plus a 26% fringe benefits add on. Applications will be accepted beginning May 1and will continue until the position is filled. Send resume and 3 letters of recommendations to: Director, Arts and Science Research Center Box RC, New Mexico State University Las Cruces, NM 88003
Hank Adams Electron Microscopy Laboratory NMSU 505-646 3600
I seem to have reached the zenith of my very limited knowledge of immunohistochemistry. I would be grateful for any assistance you can give on the following problem:
I am trying to use a fish monoclonal anitbody to vitellogenin (a yolk protein precursor) to stain fish gonad and liver 7u parafin sections. On vitellogenic oocytes I am seeing what appears to be good specific staining. The problem is that I also see the same intensity of staining with secondary only, thus my problem (TRITC goat anti-mouse; I tried FITC but got a lot of autoflouresence). I am blocking with 1% BSA 10 min. Primary incubation is overnight at room temp, concentration of 25 ug/mL; secondary is 30 min. concentration of 1:50 (also tried 1:200 same response).
Diana Papoulias Fish Biologist, Research Midwest Science Center Biological Resource Division U.S. Geological Survey 4200 New Haven Rd. Columbia, MO 65201 Tel 573 875 5399 xt 1902 Fax 573 876 1896 email: Diana_Papoulias-at-nbs.gov
Hi Paula, Dr. Beverley Giamarra has done a lot of work with HMDS. Below is a bibliography of HMDS work I am familiar with: If you hear of any others I'd appreciate your forwading them to me.
HMDS
Adams JL, Battjes DA, and Buthala DA. (1987). Biological Specimen Preparation for SEM by a Method Other Than Critical Point Drying. Proc. EMSA. 45, 956-957.
Giammara B, Baker R, Burkes E, Malangoni M, Evers B, and Hanker J. (1987a). Hexamethyldisilazane for Rapid Scanning Electron Microscopic Examination of Implant Specimens. (abs) J. Dent. Res. 66, 187.
Giammara B, DeVries W, Baker R, Dobbins J, and Hanker J. (1987b). Hexamethyldisilazane Drying for Rapid Detection of Bacteria in Implant Specimens. Proc. EMSA. 45, 878-879.
Giammara B and Hanker J. (1988a). Ruthenium Red-Osmium Bridging with TCH: New Technique to Stain Biological Specimens for Light and TEM and to Coat for SEM. Proc. EMSA. 46, 20-21.
Giammara BL and Hanker JS. (1988b). New Ruthenium Red Bridging Technique with Thiocarbohydrazide to Stain Polyanionic Biomacromolecules for Light and Electron Microscopy and to Coat Biological Specimens for Scanning Electron Microscopy. Proc. 9th Eur. Cong. Elec. Mic.
Giammara B, Washburn M, Malangoni M, Evers B, Baker R, Burkes EJ, and Hanker J. (1987c). Rapid Scanning Electron Microscopic Examination of Implant Specimens with Hexamethyldisilazane Drying. Soc. for Biomaterials Trans. X, 103.
Lamoreaux W. (1988). Prevention of Outgassing When Coating Tissues Dried with Hexamethyldisilazane (HMDS). EMSA Bull. 18, 91.
Nation JL. (1983). A New Method Using Hexamethyldisilazane for Preparation of Soft Insect Tissue for Scanning Electron Microscopy. Stain Technol. 58, 347-351.
HMDS SAFETY NOTES HMDS is flammable, use in a flammables rated exhaust hood. Avoid breathing vapors. It is toxic by inhalation, toxic sin contact with skin, or if swallowed. I have observed that HMDS seems to produce an ammonia compound when mixed with ethanol.
Other than the above common sense precautions, it is very easy to use as a final solvent treatment after fixation, post-fixation and dehydration. I have not used it myself with plant material.
good luck
Edward J. Basgall, PhD The Pennsylvania State University Surface Chemistry Group ejb11-at-psu.edu Materials Research Institute Building Ph: 814-865-0493 University Park, PA 16802-7003 FAX: 814-863-0618 } } Dear smart people, } } } I am still having problems with my cpd efforts giving me 'raisins' } especially on young plant tissue. Has anyone used HMDS? How do the } results compare to CPD, or Freon (I know I can't use freon, but it used to } be the standard)? } } Are there any tricks I should know before embarking on a HMDS experiment? } } Thanks for letting me pick your brains. } } } Paula = ) } } Paula Sicurello } UC Berkeley } Electron Microscope Lab } psic-at-uclink4.berkeley.edu
Marylin, From your message it wasn't clear to me how small your grids need to be. But here are a few suggestions anyway. Bellco Glass sells a coverslip with an etched grid pattern. I've used these very successfully for light microscopy e.g. locating cell which I have microinjected. I have also made my own indicator coverslips by using a locator EM grid as a mask and evaporating a layer of carbon (LM only) or gold (LM or EM). The gold layer need not be thick. The problems with metal evap are that (1) you must clean and bake the glass before and after metal deposition otherwise it might lift off when you place it in media or buffer (2) the metal will act as a neutral density filter and attenuate light, it may even burn up if you are using a laser for imaging (depends on incident intensity). You can avoid this problem by just working with cells off the metal. Hope these suggestions are helpful. If you have any other questions don't hesitate to contact me. Victoria Centonze Frohlich Deputy Director, IMR University of Wisconsin, Madison Symposium ($30) and Workshop ($230) on "Applications of Multiphoton Excitation Imaging" August 9 and 10, 1997 Sheraton City Center, Cleveland, OH To Register Contact: dvolkman-at-students.wisc.edu www.bocklabs.wisc.edu/imr.html
} We are looking for slides imprinted with grid squares so we can maintain } orientation/coordinates. I was wondering if anyone has knowledge of any } source that carries slides marked for orientation. Any help will be } appreciated.
I am interested in finding out some of the ways that people tend to quantify retroviral particles (as a whole, not just infectious). We have been using a vendor that performs thin-section TEM and have looked some into negative stain TEM. We have not seen consistent results from the thin section determinations, and have been working on the sample prep end to get a more consistent pellet volume and consistency to submit for TEM.
I am interested in hearing what other people tend to do for retroviral quantitation (particularly in the Biotech industry because of FDA concerns). If I could get some input from this listserv group, I would appreciate it. The address above is correct. Thank you.
Do you have any protocols for EM in-situ using digoxygenin labelled probes? My samples are already embedded in LR White, is this OK for that? These probes work really well for light microscopy in-situs.
Thanks for any information you can give me on this subject.
Paula = )
Paula Sicurello UC Berkeley Electron Microscope Lab psic-at-uclink4.berkeley.edu
From my field experience - indeed most part of stage drift in HVTEM comes from thermal influences from cooling systems. Once investigating such extensive drift by CM-20 we measured very precisely obj. lens-water temperature and found out, that the image drift observed by second highest mangnification step follows exactly the water cooler regulation. In fact all coolers which have temp. level regulation mechanism with histeresis are not suitable for todays HVTEM. Philips f.e. recommends the units with follow-up temp. regulation so theoretically (and very close practically) such unit by constant operating conditions (constant power) can reach a steady state - constant water temperature. The ,,histeresis'' units can't. When experimenting with such water supply in a/m example we achieved finally - to have the same feature in the screen center (by second highest magn.) for 30 min without any noticable movement (!) (only waht was noticable - growing contamination ring). That was in very patient test conditions - it's not the case by normal work, ofcourse. regards Krzysztof M. Herman E.Eng M.Sc. Philips El.Opt. Service Specialist LabSoft S.c. 05-500 Piaseczno, 21 Kosciuszki Str. tel/fx: (48 22) 7502024, 7502028, 7570671 fax only: (48 22) 483787, Email: labsoft-at-ikp.atm.com.pl
I once again have called on the services of Bernie Kestel from Argonne National Laboratory and he has given me the following information:
"Selective etch of Cu from cobalt , non-electolytic. Try dilute nitric acid in methanol, NOT ethyl alcohol, (an explosive mixture). Water as a solvent may also work. Try 3-4 % acid & increase concentration if needed to get some action. Heat is generated when mixing the acid mixture, so cool with ice or dry ice before adding acid. Use at room temp. Cobalt is usually electropolished with perchloric acid mixtures so the nitric acid may not attack it much. In electropolishing, Cu works in nitric solutions while Co works in perchloric solutions. Even my non-acid bath, BK-2, does not work on Cu. I don't know how to thin Cu & Co at the same rate. Develop a new non-acid electrolyte, perhaps."
Bernie does his work with our Model 550 Single Vertical Jet Electropolisher, but other jet polishers will certainly work well too.
You can get additional information on the Jet Polisher (Now Model 550D) on our web site at http://www.southbaytech.com.
DISCLAIMER: As we do manufacture the Model 550D Single Vertical Jet Electropolisher, I obviously have a vested interest in promoting its use.
David Henriks TEL: 800-728-2233 (toll-free in USA) South Bay Technology, Inc. 714-492-2600 1120 Via Callejon FAX: 714-492-1499 San Clemente, CA 92673 USA e-mail: henriks-at-southbaytech.com
Dear Diana, Have you tried blocking with 10% goat serum? I also add 10% goat serum in my primary and secondary antibodies. Another point to consider is your source of secondary! Is the staining specific or non specific...does it stain up the same area of what you expect to get with the addition of primary? I would suggest you try some other tissues as positive and negative controls and if all else fails, maybe try another source of secondary antibody...or if possible, purify the secondary through an appropriate affinity column. Hope this helps!
K.M.Khoo, Department of Biochemistry, Faculty of Medicine, National University of Singapore.
On Thu, 24 Apr 1997, Diana Papoulias wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } List readers, } } I seem to have reached the zenith of my very limited knowledge of } immunohistochemistry. I would be grateful for any assistance you can } give on the following problem: } } I am trying to use a fish monoclonal anitbody to vitellogenin (a yolk } protein precursor) to stain fish gonad and liver 7u parafin sections. } On vitellogenic oocytes I am seeing what appears to be good specific } staining. The problem is that I also see the same intensity of } staining with secondary only, thus my problem (TRITC goat anti-mouse; } I tried FITC but got a lot of autoflouresence). I am blocking with 1% } BSA 10 min. Primary incubation is overnight at room temp, } concentration of 25 ug/mL; secondary is 30 min. concentration of 1:50 } (also tried 1:200 same response). } } } Diana Papoulias } Fish Biologist, Research } Midwest Science Center } Biological Resource Division } U.S. Geological Survey } 4200 New Haven Rd. } Columbia, MO 65201 } Tel 573 875 5399 xt 1902 } Fax 573 876 1896 } email: Diana_Papoulias-at-nbs.gov } }
This is THE FINAL URGENT REQUEST for your comments, suggestions, etc about the 3RD RFD for my PROPOSED NEW NEWSGROUP SPECIALIZING IN IMMUNOCYTOCHEMISTRY/IMMUNOHISTOCHEMISTRY/OTHER RELATED AFFININITY METHODS called "sci.bio.immunocytochem" now posted in "news.groups"
The latter contains material of a rather varied nature (!) but it is necessary to use this unmoderated group to hold the discussions about all the different proposed new groups.
So, if you are connected to Usenet, and you are keen to see A NEW IMMUNOCYTOCHEMISTRY NEWSGROUP, then please go to "news.groups", ignore all the rubbish, look for articles posted on 20.4.97 or 24.4.97 (or later) and you should find one of my postings "3RD RFD: sci.bio.immunocytochem". Select "follow-up article" (or equivalent) from your newsreader menu, and post your message so that it appears under the 3RD RFD.
If you don't have access to Usenet, you can read the proposal at the Royal Microscope Society web site {http://www.rms.org.uk} , or at the Introduction to Immunocytochemistry (Center for Cell Imaging Department of Cell Biology Yale University School of Medicine) web site {http://info.med.yale.edu/cellimg/CCIimmuno.html}
MANY THANKS to all of you who have already posted your responses to my RFDs! BUT WE STILL NEED MORE DISCUSSION in news.groups please! SOON I shall be posting my "CALL FOR VOTES" to the people in charge of Usenet newsgroups. When you see "CFV: sci.bio.immunocytochem", you will be able to e-mail your vote to the vote-taker.
Hi: I have received two E-mail letters from Philip Koeck over the past week or so through this forum. Both letters I can not read or delete. Whenever I try I get an error message saying that the program has performed an illigal action and turns netscape off. Anybody having the same problem or any solutions? Thank you, Peter Jordan
Peter Jordan wrote: } } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Hi: } I have received two E-mail letters from Philip Koeck over the past week } or so through this forum. Both letters I can not read or delete. } Whenever I try I get an error message saying that the program has } performed an illigal action and turns netscape off. Anybody having the } same problem or any solutions? Thank you, Peter Jordan
Yes I am. It's very frustrating. I'd appreciate any help.
Peter Jordan wrote: } } Hi: } I have received two E-mail letters from Philip Koeck over the past week } or so through this forum. Both letters I can not read or delete. } Whenever I try I get an error message saying that the program has } performed an illigal action and turns netscape off. Anybody having the } same problem or any solutions? Thank you, Peter Jordan .......................... I also use Netscape 3.0 and have the same problem. I don't have a solution, but I do have a way to get rid of the "poison message":
(1) read, and delete or transfer all the rest of the mail in the Inbox, but don't try to read or delete the "problem" message. (This is tricky, and you may have to "error-out" and reload Netscape a couple times.) (2) When the Inbox is empty except for this message, exit from Netscape. (3) Go to the NETSCAPE\NAVIGATOR\MAIL subdirectory, and locate the file "Inbox". Delete it (or rename it if you want to be cautious). (4) The next time you load Netscape, it will recreate an empty Inbox file.
This is ugly, but it's the only way I've found to get rid of the darn thing!
I hope somebody who doesn't have this problem will help us out by sending a message to the originator to find out what he is doing! I can't open his messages to get a return address. -- Fred Schamber ....................... mailto:fhscham-at-SGI.NET
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Frederick Schamber wrote:
I also use Netscape 3.0 and have the same problem. I don't have a solution, but I do have a way to get rid of the "poison message":
(1) read, and delete or transfer all the rest of the mail in the Inbox, but don't try to read or delete the "problem" message. (This is tricky, and you may have to "error-out" and reload Netscape a couple times.) (2) When the Inbox is empty except for this message, exit from Netscape. (3) Go to the NETSCAPE\NAVIGATOR\MAIL subdirectory, and locate the file "Inbox". Delete it (or rename it if you want to be cautious). (4) The next time you load Netscape, it will recreate an empty Inbox file.
This is ugly, but it's the only way I've found to get rid of the darn thing!
I use Netscape V. 3.01 and I have the same problem. This is the way I deal with it:
I discover the message that is causing trouble (sometimes you'll have to launch Netscape several times to find out the message). Then I mark a group of messages including the "problem" message (never mark it at first or last. Keep it among others). You'll have to do the same again when they are transferred to the trash directory. This procedure works well most of time.
I would like any information you have on the KEVEX 8000 upgrade system. I am specifically looking for how many channels can be imaged at once. i.e. number of EDS maps displayed at one time and how many external channels (WDS) can also be displayed with the EDS?
Also, how do you like the operation of the system?
The problem appears to be associated with Netscape Email readers as I can read and delete the file using other Email programs (i.e standard Mail on Unix and Eudora on PC or Mac Platforms). I have sent a message both to NETSCAPE and to Philip Koeck {Philip.Koeck-at-csb.ki.se} the originator of the message. I noticed the Koeck uses Netscape Gold V 3.01 on a Windows 95 Machine (you can find this out by reading his Email header file)
It would probably help for those of you running Netscape who have the problem to also report it to NETSCAPE. They have an on-line form for this at:
To all who responded to my plea for help: Thank you very much. Sandwiching the "bad" files between two "good" files allows you to move them into the trash bin and then to delete them. For all computer greenhorns like me this is done by clicking on the good file, then holding down the shift key clicking on the other good file. All files between the clicked files will be marked and then can be moved. It was realy nice to get all these respones. Thanx again, Peter Jordan
Hi: I forgot to give credit to Dan Kaszubski for the solution on how to delete Philip Koeck's E-mail letters. Thank you, Dan. I did not realize how many of us had this problem. The recipe is in my other letter. Peter Jordan
Dear all, I am aware of a number of programs that can produce calculated diffraction patterns given the unit cell parameters and the location of all atoms in the unit cell. This poses difficulties for some materials with more complex structures, however, where site occupancies are not 100% and where the 'unit cell' is somewhat of an average structure (this occurs for some oxides). This can be overcome if a good crystal structure refinement has been done from XRD where average atomic positions and site occupancies have been determined. Many structures have not been studied in this detail, however, and the only information that is available is the unit cell parameters and the space group. Is there any software that can plot electron diffraction patterns from this limited information.
I hope there is someone who can help me with this.
++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ Ian MacLaren, Tel: (44) (0) 121 414 3447 IRC in Materials for FAX: (44) (0) 121 414 3441 High Performance Applications, email: I.MacLaren-at-bham.ac.uk The University of Birmingham, http://web.bham.ac.uk/I.MacLaren/ Birmingham B15 2TT, England. ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++
} } } } } } } } } } } } } } } } } } } } } .. } } I am a research scientist in Columbia University and I need some Quartz } Cover Slips badly for one of my projects. I was unable to locate any. } Would you please kindly send me a list of vendors who may carry this } product that I can buy from. } My email address is hl51-at-columbia.edu } I would appreciate it very much, } } Sinserely your, } } Hui Lao, M.D. } } ******************************************************* G.W. Erdos, Ph.D. Phone: 352-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax: 352-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 http://www.biotech.ufl.edu/~emcl/ Home of the #1 Gators ***** "Many shall run to and fro, and knowledge shall be increased" Daniel 12:4
Dear Marilyn, Following is information on the grided slide. FIELD FINDER Microslide Field Finder helps you index points of interest on microscope slide the same way you find them on a road map. Standard 75X25mm slide has photographically imprinted grid of 1mm squares, subdivided into 0.1mm intervals. eash square is marked with letter and number. You center detail in field of microscope...replace specimen slide with field finder...and read coordinates. The slide is provided by the following: Fisher Scientific 1-800-766-7000 Catalogue number# 12-454 Price each $142.00
I am not affliated with this company in any way. I am just passing on the information.
Dear list, We are looking for slides imprinted with grid squares so we can maintain orientation/coordinates. I was wondering if anyone has knowledge of any source that carries slides marked for orientation. Any help will be appreciated. with regards,
************************* * Marilyn Wadsworth * * mwadswor-at-zoo.uvm.edu * *************************
I am Electrical-Engineer , and i have in propiety Electron -microprobe i I would like any information you about electron -microprobe result of smectite Clinoptilolites and others Zeolites .
I think you will probably get lots of replys from the vendors for your question, but I thought I'd tell you a few of the companies that I have used for years with satisfactory results.
Electron Microscopy Sciences, 321 Morris Rd, Box 251, Ft. Washington, PA 19034, 1-800-523-5874
Ernest F. Fullam, Inc., 900 Albany Shaker Rd, Latham, NY 12110, (518)785-5533
Ted Pella, Inc., e-mail tedpel-at-aol.com or tedpel-at-snowcrest.net
These are just my own picks, I'm not affiliated with any of them.
Karen Pawlowski
On Fri, 25 Apr 1997, Sandi Burany wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } } Hello Suppliers: } } Would you please send me the catalogs of the materials needed for TEM, } SEM experiments. } } Thank you in advance. } } Sandy Burany } CHIPWORKS } 3685 Richmond Rd, Suite 500 } Ottawa, ON K2H 5B7 } Canada } (613) 829-0414 Ext. 3056 } FAX: (613) 829-0515 } Email:sburany-at-chipworks.com }
Yves MANIETTE wrote: } I am interested in any hint in order to perform the calibration of a TEM } high temperature sample holder test, up to 1000 degrees C. The idea is } looking at various sample with a known structural transformation ocurring } at precise temperature, and compare this "real" temperature with the one } given by the sample holder thermocouple=8A
Be cautious=8A
We have often observed that phase transition in perovskites and memory alloys (in the range of 100=B0C to -200=B0C) did occur at significant lower temperature in TEM thin foils than in bulk material (some tens of =B0C). We do not believe that it is due to errors in the temperature calibration, nor temperature gradient in the sample/sample holder, or to a heating effect by the electron beam. The source of this effect has probably to beaccounted for in the reduced size of the sample (=892D phenomena) or the sample surfac= e quality (presence of an amorphous layer on top/bottom of ion milled samples for example).
Remember that 2.5nm gold particles suffer fron a "size effect" and melt below 300=B0C instead of more than 1000=B0C for the bulk!
Remember also that the beam may significantly increase the temperature of particles deposited on a film when they start to be absorbant (thick observable samples). We vaporized micrometers sized silver spheres deposited on a carbon film just by focusing the illumination beam (100 kV, W filament, largest standard condenser aperture)! But very small spheres were not significanltly heated because of their high surface/volume ratio more favourable to cooling by radiation than heating by absorption.
Good luck Philippe A. Buffat
__________________________________________________________________ Philippe Buffat Ecole Polytechnique Federale de Lausanne (EPFL) Centre Interdepartemental de Microscopie Electronique Address: EPFL-CIME, Batiment MX-C, CH-1015 Lausanne, Switzerland Phone: +41(21)693 29 83 Fax: +41(21)693 44 01 (Central European Time) E-mail: philippe.buffat-at-cime.uhd.epfl.ch, WWW URL http://cimewww.epfl.ch/ ______________________________ Eudora F2.1 ___________________________
I do regurlarly count of Retrovirus particles in negative staining for companies in the Biotech industry. Concerning count in thin sections, I tried, but I abdict because there were too much parameters giving false results: size of the pellet, type of grids, thickness of the sections, etc. Thin sections are good for know how many cells are infected and to determine clearly the type of Retrovirus particles. I have good results in negative staining. I made count in mixing supernatant containing Retrovirus particles with latex beads of known concentration. I ultracentrifuge the mix on a grid in a Beckman Airfuge at 20-30 psi for 5 minutes, and stain with PTA 3%, pH 6. This count in negative staining is more reliable and less expensive than thin sections. If you need more details, do not hesitate to contact me.
Robert Alain Institut Armand-Frappier Laval, Quebec tel: 514-687-5010 ext 4388
Robert_alain-at-IAF.uquebec.ca
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I am interested in finding out some of the ways that people tend to quantify retroviral particles (as a whole, not just infectious). We have been using a vendor that performs thin-section TEM and have looked some into negative stain TEM. We have not seen consistent results from the thin section determinations, and have been working on the sample prep end to get a more consistent pellet volume and consistency to submit for TEM.
I am interested in hearing what other people tend to do for retroviral quantitation (particularly in the Biotech industry because of FDA concerns). If I could get some input from this listserv group, I would appreciate it. The address above is correct. Thank you.
new to the field of EM polymer blends I would like to ask you for the best way to prepare PE/PS blends for structure determination with TEM.
Of course the material has to be stained and cut, but:
- what is the best staining agent (OsO4, RuO4, or something else)? - is it better, first to stain and then to cut or the other way round? - which temperature is the best for cutting, liquid nitrogen or room temperature?
I would appreciate ever tip & trick you can give on handling this kind of material.
Tanks in advance
Petra
-------------------------------------------------------------- Dr. Petra Wahlbring Centre de Recherche Public Centre Universitaire (CRP-CU) Laboratoire d'Analyse des Materiaux (LAM) 162a, av. de la Faiencerie L-1511 Luxembourg tel. +352-466644-402 fax +352-466644-400 e-mail: petra.wahlbring-at-crpcu.lu or 100112.2335-at-compuserve.com
Use our Web page, "LabNet" for a review of 89 products. it is http://www.shore.net/~catalogs
Regards, Elinor Solit Director of Publications The Microscope Book
On Fri, 25 Apr 1997, Sandi Burany wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } } Hello Suppliers: } } Would you please send me the catalogs of the materials needed for TEM, } SEM experiments. } } Thank you in advance. } } Sandy Burany } CHIPWORKS } 3685 Richmond Rd, Suite 500 } Ottawa, ON K2H 5B7 } Canada } (613) 829-0414 Ext. 3056 } FAX: (613) 829-0515 } Email:sburany-at-chipworks.com }
Dear All, I have a student doing Mossbauer and EM studies of commercial ferrosilicons. We have a need to check our microanalysis results with regard to Fe and Si in both the SEM and TEM. Does anyone have or know where we can get Fe2Si and Fe3Si (or if all else fails anything close) to make standards? Thanks Mike
Michael J Witcomb PhD Electron Microscope Unit University of the Witwatersrand Private Bag 3 WITS 2050 South Africa
We are contracting our topical issues for 1998, and have resolved all problems of the huge backlog that plagued us in 1995 and 1996. If you are interested in serving as a Guest Editor for a particular topic, please e-mail me direct and present your idea. There is an honorarium. All articles will be published within about 6 months after they are received by the publisher in New York, so hot research articles would fit in nicely.
John E. Johnson, Jr., Ph.D. Editor, Microscopy Research and Technique sdinfo-at-worldnet.att.net
NYMA Inc., an aerospace engineering company serving NASA at Lewis Research Center, is seeking to fill the following two challenging positions;
SCANNING ELECTRON MICROSCOPIST
Required to operate and maintain three scanning electron microscopes, and provide materials characterization support. The individual must also be able to assist and instruct research staff in the principles and operation of scanning electron microscopy.
Requirements: B.S. degree with 3 years experience or 7+ years of extensive hands on experience in electron microscopy. Must have a thorough working knowledge of x-ray energy/wavelength dispersive spectroscopy, preparation equipment, and vacuum systems. A working knowledge of x-ray diffraction and electronics is a plus. Excellent communication and interpersonal skills are required.
TRANSMISSION ELECTRON MICROSCOPIST
Extensive experience in operating, and maintaining a 200 KeV transmission electron microscope in support of material characterization of advanced high temperature materials. The successful candidate will work with materials researchers to understand materials' properties.
Requirements: Ph.D. in material science with 3+ years extensive experience operating a transmission electron microscope using SAED, CBED, XEDS, EELS, and PEELS to analyze a wide range of materials. The individual we seek must have extensive experience in sample preparation using electro-polishing, ion-milling, PIPS, and PIMS. A working knowledge of a dedicated STEM and x-ray diffraction is a plus. Excellent communication and interpersonal skills are required.
Qualified candidates should submit a resume to : Todd Leonhardt, NYMA, Inc., Mail Stop 105-1, 2001 Aerospace Parkway, Brook Park, OH 44142 or by E-mail to todd.a.leonhardt-at-lerc.nasa.gov
Posted Date: April 28,1997 Closing Date: Open until filled
With PE/PS I have gotten good results if I first cut and then stain. I do use cryo sectioning and I cool the sample to about -100C (room temperature will not work because the Tg of PE is too low). I use folding grids to collect my sections (dry) and then I stain with RuO4 which will stain the PS. You can also stain the block first and then cut but then you are limited to sections near the surface of the specimen (the stain does not penetrate too far). Even then, you might need to re-stain your sections. Finally, I deposit carbon.
I hope this helps,
Jordi Marti ----------------------------------
Hello polymer experts,
new to the field of EM polymer blends I would like to ask you for the best way to prepare PE/PS blends for structure determination with TEM.
Of course the material has to be stained and cut, but:
- what is the best staining agent (OsO4, RuO4, or something else)? - is it better, first to stain and then to cut or the other way round? - which temperature is the best for cutting, liquid nitrogen or room temperature?
I would appreciate ever tip & trick you can give on handling this kind of material.
Tanks in advance
Petra
-------------------------------------------------------------- Dr. Petra Wahlbring Centre de Recherche Public Centre Universitaire (CRP-CU) Laboratoire d'Analyse des Materiaux (LAM) 162a, av. de la Faiencerie L-1511 Luxembourg tel. +352-466644-402 fax +352-466644-400 e-mail: petra.wahlbring-at-crpcu.lu or 100112.2335-at-compuserve.com
I have two questions about type of oil you use for mechanical and diffusion pumps for the microscopes.
(I) We have a JEOL SM 35-C (SEM) since many years ago. In the last two years we have often seen oil drops condensed on the EDS detector, inside the chamber (at least it looks like oil, light green-dark yellow coloured drops). We've been using Dow Corning 704 silicon oil for the diffusion pump since ever. Now, we're not sure what it is going on, we suspect either: * backstreaming problems from the mechanical pumps oil * not sufficiently high vapor pressure for the DC 704
What do you think about it? any experience?
(II) We have been using Edwards 15 oil for the mechanical pumps. This time we have gotten a very high quotation (according to our reduced budget) for this oil. We're thinking of looking for other brand of the same type to replace it. Any idea what to buy?
We would appreciate to hear any suggestion from any of you.
Many thanks in advance. Sincerely,
Silvia Montoro Centro Regional de Investigacion y Desarrollo Guemes 3450 Santa Fe - Argentina csedax-at-arcride.edu.ar
NYMA Inc., an aerospace engineering company serving NASA at Lewis Research Center, is seeking to fill the following two challenging positions;
SCANNING ELECTRON MICROSCOPIST
Required to operate and maintain three scanning electron microscopes, and provide materials characterization support. The individual must also be able to assist and instruct research staff in the principles and operation of scanning electron microscopy.
Requirements: B.S. degree with 3 years experience or 7+ years of extensive hands on experience in electron microscopy. Must have a thorough working knowledge of x-ray energy/wavelength dispersive spectroscopy, preparation equipment, and vacuum systems. A working knowledge of x-ray diffraction and electronics is a plus. Excellent communication and interpersonal skills are required.
TRANSMISSION ELECTRON MICROSCOPIST
Extensive experience in operating, and maintaining a 200 KeV transmission electron microscope in support of material characterization of advanced high temperature materials. The successful candidate will work with materials researchers to understand materials' properties.
Requirements: Ph.D. in material science with 3+ years extensive experience operating a transmission electron microscope using SAED, CBED, XEDS, EELS, and PEELS to analyze a wide range of materials. The individual we seek must have extensive experience in sample preparation using electro-polishing, ion-milling, PIPS, and PIMS. A working knowledge of a dedicated STEM and x-ray diffraction is a plus. Excellent communication and interpersonal skills are required.
Qualified candidates should submit a resume to : Todd Leonhardt, NYMA, Inc., Mail Stop 105-1, 2001 Aerospace Parkway, Brook Park, OH 44142 or by E-mail to todd.a.leonhardt-at-lerc.nasa.gov
Posted Date: April 28,1997 Closing Date: Open until filled
I am interested in studying silver deposited onto Pt(111) by STM in electrochemical conditions.Does anybody tried (and succed) to obtain EC-STM images with this system ? I didn't find any publication on this subject. I know that there are allready publications with this system studied in UHV but my results are far away from what they published.
Thank you very much for your cooperation. ------------------------------------------ :-) :-) :-) :-) :-) :-) :-) :-) :-) :-) :-) :-) ------------------------------------------ Lorena H. Klein Laboratoire de Physico-Chimie des Surfaces (CNRS-URA 425) 1, Pl. A. Briand F-92195 Meudon FRANCE tel: +33 1 45075361, fax: +33 1 45075858 e-mail: lorena.klein-at-cnrs-bellevue.fr
I have a good reference that you might want to check out:
"Ruthenium Tetraoxide Staining of Polymers for Electron Microscopy", by J.S. Trent, et al, Macromolecules Vol. 16, #4, pp. 589- 1983.
This article is VERY informative.
Regards,
Bob *********************************** Bob Citron Chiron Vision 555 W. Arrow Hwy Claremont, CA 91711 ph: (909)399-1311 email: Bob_Citron-at-cc.chiron.com ***********************************
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Hello polymer experts,
new to the field of EM polymer blends I would like to ask you for the best way to prepare PE/PS blends for structure determination with TEM.
Of course the material has to be stained and cut, but:
- what is the best staining agent (OsO4, RuO4, or something else)? - is it better, first to stain and then to cut or the other way round? - which temperature is the best for cutting, liquid nitrogen or room temperature?
I would appreciate ever tip & trick you can give on handling this kind of material.
Tanks in advance
Petra
-------------------------------------------------------------- Dr. Petra Wahlbring Centre de Recherche Public Centre Universitaire (CRP-CU) Laboratoire d'Analyse des Materiaux (LAM) 162a, av. de la Faiencerie L-1511 Luxembourg tel. +352-466644-402 fax +352-466644-400 e-mail: petra.wahlbring-at-crpcu.lu or 100112.2335-at-compuserve.com
Message-Id: {1.5.4.32.19970428230321.0068b974-at-pop3.unsw.edu.au} X-Sender: s7001031-at-pop3.unsw.edu.au X-Mailer: Windows Eudora Light Version 1.5.4 (32) Mime-Version: 1.0 Content-Type: text/plain; charset="us-ascii"
} -------------------------------------------------------------- } Dr. Petra Wahlbring } Centre de Recherche Public Centre Universitaire (CRP-CU) } Laboratoire d'Analyse des Materiaux (LAM) } 162a, av. de la Faiencerie L-1511 Luxembourg } tel. +352-466644-402 fax +352-466644-400 } e-mail: petra.wahlbring-at-crpcu.lu or 100112.2335-at-compuserve.com } } Dear Petra,
The book you must read is POLYMER MICROSCOPY by Linda Sawyer and David Grubb Chapman & Hall 2nd edition 1996 ISBN 0 412 60490 6
Try Chapman & Hall 2-6 Boundary Row, London SE1 8HN UK
one of the problems associated with lead citrate stain preparation is the presence of carbonate in the NaOH. John Luft came up with a solution for this years ago. Here is a copy of a handout from his 1976 seminar series that covers both uranyl acetate and lead citrate stain preparation.
steve
{bold} {fontfamily} {param} Times {/param} {bigger} {bigger}
NOTES ON URANYL AND LEAD STAINING (J.H. Luft) {/bigger} {/bigger} {/fontfamily} {/bold} {fontfamily} {param} Times {/param} {= bigger} {bigger}
{bold} {underline} Uranyl Acetate
{/underline} {/bold}
A saturated solution of uranyl acetate in distilled water is commonly used to stain EM sections either alone or preceding a lead stain.=20 However, uranyl acetate is not a well-behaved salt either as crystals in the bottle or as a stock solution. It tends to hydrolyse to form the basic acetate, U02(OAc) 2.2H20 + H20 --} U02(0H) (0Ac) + H0Ac and free acetic acid which produces the smell of acetic acid in a bottle of crystals. The easiest way to detect the presence of the basic acetate is to dissolve some of the uranyl acetate in question; at room temperature, it should dissolve (overnight) to give a crystal clear 8% solution. Any pale yellow residue on the bottom is the basic acetate, and the solution made up with this uranyl acetate will be saturated with the basic acetate. This basic acetate probably is the main source of contamination of EM sections from uranyl staining. Even originally completely clear uranyl acetate solutions may begin to decompose to precipitate out the basic acetate, but it is erratic - a week, or a year.
Large crystals of uranyl acetate seem to be much more stable against hydrolysis form moisture in the air than broken crystals. Powdered uranyl acetate is the worst offender in this regard, although {underline} fresh {/underline} powdered material may be satisfactory. In the past, uranyl acetate from Fisher Chemical Company has been the best because it is supplied as relatively large crystals. If no source of easily soluble uranyl acetate can be obtained, it is possible to recrystallize good material from even a badly hydrolysed supply as follows:
About 50 gm of the decomposed uranyl acetate is added to 100 ml of 10% acetic acid and heated to 70-80 {/bigger} {/bigger} {/fontfamily} {bigger} {bigger} {fontfamily} {param} MS_Li= neDraw {/param} =DF {/fontfamily} {fontfamily} {param} Times {/param} C on a hot plate with stirring for 10-15 minutes. the hot solution is then filtered through a conical paper filter into a clean, covered beaker and allowed to cool with frequent stirring, otherwise the crystals form on the sides and bottom of the beaker). It can be cooled further with ice for a higher yield. After several hours, the crystals can be filtered and sucked dry by vacuum for an hour on a Buchner funnel and stored in a tightly capped bottle. The residue from the original 50 gms. can be added to more old uranyl acetate and another batch recrystallized. This recrystallized uranyl acetate should give clear solutions for several years.
{bold} {underline} Lead Stains
{/underline} {/bold}
Reynolds alkaline lead citrate (E.S. Reynolds, J. Cell. Biol. {underline} 17 {/underline} , 208, 1963) is one of the most successful of the alkaline lead stains because of its intensity and relative freedom from contamination. Nevertheless, problems do arise with it, and contamination may appear mysteriously. Most of the trouble, I believe, is due to the problems of storing strongly alkaline solutions. These=20 rapidly attack and dissolve silicate from glass bottles (including Pyrex), and pick up C02 from the air if they are stored in polyethylene bottles. Solutions of Reynolds stain stored in small polyethylene bottles deteriorate and develop crystals or a residue on the side of the bottle in several months. It can be shown that this is a result of the C02 absorption, because a similar polyethylene bottle of stain kept {underline} within {/underline} a larger, wide-mouth, bakelite-capped bottle, which contains a little moist barium hydroxide as a C02-trap, remains clear and useful for several years.
The crystals are not a direct result of carbonate accumulation itself, because sodium carbonate can be dissolved directly in Reynolds stain without precipitation. Instead, the absorbed C02 acting as an acid probably reduces the pH of the stain to the point that the lead complex precipitates, and where the stain then does become sensitive to carbonate precipitation.
The nesting of a polyethylene bottle inside a glass bottle combines the alkali-resistance of polyethylene with the C02-impermeability of glass. This is an ideal solution for storage of alkaline materials but the solutions should be C02-free when they are made up as well. Reynolds advises the use of freshly boiled distilled water for this reason, and this works well. (Many stills can actually {underline} concentrate {/underline} C02 in the distillage, and the C02-content of some tap water varies from winter to summer, so boiling is a useful habit). It is satisfactory to boil a liter of distilled water in an Erlenmeyer flask down to 600-700 ml, then cover with a beaker and cool in tap water and use within a few hours.
Boiling water and waiting each time to make Reynolds stain is a nuisance, and it would be useful to make up stock solutions once with boiled distilled water and have it on hand thereafter. This is the recipe:
{bold} Solution A {/bold} : Lead nitrate, Pb(N03)29 31.67 gm made up to 500 ml with boiled, distilled water, plus 10 drops concentrated nitric acid.
{bold} Solution B: {/bold} Sodium citrate, Na3 C6H507.2H209 41.9 gm made up to 500 ml with boiled, distilled water. Add 5 drops of {underline} solution A {/underline} as a preservative. Both can be stored in regular capped glass bottles. The stain is made by adding 21 ml of {underline} solution A {/underline} to the 50 ml clean polyethylene bottle, using a 25 ml graduate. Rinse the graduate with distilled water and then add 21 ml of {underline} solution B {/underline} , and swirl to mix. Add 8 ml of 1.0N Na0H and again swirl. The solution clears and produces 50 ml of Reynolds lead stain ready for use.
The source of the 1.0N Na0H is also a problem, since C02 adds rapidly to pellets of Na0H or to a stock solution. (10 N Na0H is available "carbonate-free" in polyethylene bottles. It may have been carbonate-free when it was made, but there is plenty of carbonate in it when it arrives). A neat device for ready access to carbonate-free Na0H at any time is the following:
Take a clean 250 ml polyethylene bottle and fill it about half full of Na0H pellets (from a freshly opened bottle, but this is not essential). Then fill it about 3/4 full with distilled water. Quickly cap and submerge in cold running tap water because much heat is generated.=20 Shake for an hour or two and then allow to stand on the laboratory shelf for two weeks or longer. This produces saturated Na0H and a layer of transparent Na0H crystals will develop between the pellets below and the solution above. With a {underline} plastic {/underline} 20 ml syringe and large (15 ga.) needle, suck up 13.0 ml of the saturated solution and dilute to 250 ml with boiled distilled water. Store this in a polyethylene bottle enclosed in a larger wide-mouth jar capped tightly and containing moist barium hydroxide. This combination keeps the NaOH carbonate-free indefinitely. Be sure to rinse the plastic syringe several times to remove the silicone oil lubricant, or use a de-greased plastic syringe. The saturated NaOH rapidly attacks a glass syringe. This works because although the polyethylene bottle is porous to C02, the saturated Na0H precipitates out the Na2C03 which continuously collects but floats on the surface. Therefore, don't shake the saturated Na0H before using it.
A question for the Microscopy Listserver on behalf of David Grattan;
--------------------------------------------------------------- We are dealing with an antigen in brain tissue that seems to be adversely affected by paraformaldehyde, and would like to find a gentler fixative that still preserves neuronal morphology.
Has anybody tried fixation with carboiimide (1-ethyl-3-(3-dimethylaminopropyl)carodiimide). We seem to recall a reference to this as an EM fixative that also provided good morphology for immunohistochemistry, especially for neurons.
Any suggestions/ideas regarding this method of fixation would be appreciated. ------------------------------------------------------------------ Dr. David Grattan Department of Anatomy and Structural Biology School of Medical Sciences, University of Otago P.O. Box 913, Dunedin, New Zealand Ph: (64)(3) 479-7442 (office), 479-7345 (lab) Fax: (64)(3)479-7254 Email: david.grattan-at-stonebow.otago.ac.nz Homepage: http://www.otago.ac.nz/ResearchPostGrad/Neuro/Grattan _______________________________________________________________
----------------------------------------------------------------------- Richard Lander Senior Technician South Campus Electron Microscope Unit c/- Pathology Department Otago Medical School P.O. Box 913 Dunedin New Zealand. Tel. National 03 479 7301 Fax. National 03 479 7254
"Southernmost EM Unit in the World!" ------------------------------------------------------------------------
Message on behalf of Richard Easingwood; } } We are looking at the various options for large-format (4 x 5 inch) } negative scanners and wonder whether we could get some feedback from actual } users. } I have heard that the Leaf 45 scanner (which I understand was the Rolls } Royce/Cadillac of scanners) is discontinued and no longer available - can } anyone confirm this? } } Does anyone have experiences (good or bad) with the Polaroid SprintScan 45? } } We want to use the scanner for getting high quality resolution scans from } our sheet film TEM micrographs. } } Thanks in advance for any feedback. } } Regards, } } Richard } } Richard Easingwood } South Campus Electron Microscope Unit } School of Medical Sciences } University of Otago } PO Box 913 } Dunedin } NEW ZEALAND } } Telephone: 64-03-479 7301 } Facsimile: 64-03-479 7254 } } e-mail: richard.easingwood-at-stonebow.otago.ac.nz } } }
I'm sorry about the inconvenience caused by my unreadable mail. Thanks to our friendly neighbourhood SysOp I think I have found the problem (if You can read this). My postal adress somehow ended up in the field for the reply adress. If You still have an undeletable Email somewhere You can get rid of it by editing the mail file in a word processor. -- Philip Koeck Karolinska Institutet Dept. of Bioscience Novum S-14157 Huddinge Sweden Tel.: +46-8-608 91 86 Fax.: +46-8-608 92 90 Email: Philip.Koeck-at-csb.ki.se http://www_scem.csb.ki.se/pages/philip.html
I have just read the interesting post of Steve Barlow in which he gives John Luft's method for re-crystallizing Uranyl acetate to minimize stain artefact. The logic seems pretty good and I am willing to accept it. I am not all that eager, however, to run out and waste a couple hours recrytallizing a relatively toxic powder. This seems to me the perfect thing for an EM supplier to offer. They all sell UA but wouldn't it be nice if someone sold stocks of UA with certified recrystallization dates? I would be willing to pay a little extra for that. Furthermore, why doesn't any supplier sell UA in smaller aliquots in rubber stopper vials so that one could make up small amounts without having to worry about students spilling toxic dust all over the scale, etc. I know a lot of EM suppliers monitor this list and would be pleased to see them responsed.
Thomas E. Phillips, Ph.D. Associate Professor of Biological Sciences Director, Molecular Cytology Core Facility 3 Tucker Hall University of Missouri Columbia, MO 65211 (573)-882-4712 (voice) (573)-882-0123 (fax)
} } We are looking at the various options for large-format (4 x 5 inch) } } negative scanners and wonder whether we could get some feedback from actual } } users. } } I have heard that the Leaf 45 scanner (which I understand was the Rolls } } Royce/Cadillac of scanners) is discontinued and no longer available - can } } anyone confirm this? } } } } Does anyone have experiences (good or bad) with the Polaroid SprintScan 45? } } } } We want to use the scanner for getting high quality resolution scans from } } our sheet film TEM micrographs. } } } } Thanks in advance for any feedback. } } } } Regards, } } } } Richard } } } } Richard Easingwood } } South Campus Electron Microscope Unit } } School of Medical Sciences } } University of Otago } } PO Box 913 } } Dunedin } } NEW ZEALAND } } } } Telephone: 64-03-479 7301 } } Facsimile: 64-03-479 7254 } } } } e-mail: richard.easingwood-at-stonebow.otago.ac.nz } } } } } Recently there have been several postings about negative scanners. We have been using the Agfa Arcus II for the last two years with somehat satisfactory results, however, we cannot seem to scan lattice images when they are taken below 340kx. Agfa now has a new product, the Duoscan that may allow one to do this still at a reasonable price. The Arcus II is a 12 bit scanner with 600x1200 dpi optical resolution and currently sells for about $1,800. The Duoscan has a better resolution I think around 2000x2000 dpi and I believe sells for about $5,000. We are trying to evaluate it right now.
Thanks so much to all the colleagues who took the time to offer references, web sites, comments and answers to the questions for Kelly's intergrated science project. She in compiling data, reading material, visiting web sites and getting other suggested sources. The wonderful thing is that she is learning and is so enthusiastic about the scientists all over the world who know EM and were kind enough to point her in the right direction. From the wilds of Africa to the shores of France and Australia and right here in the U.S. (and many other countries), there is a bond that has formed via science, and Kelly will always remember that people cared. What wonderful mentors you have been!!
Thank you all sincerely, Linda M. Fox lfox1-at-wpo.it.luc.edu
At 04:46 PM 4/29/97 +1200, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Yes, this is true. However, you may be able to locate a refurbished Leaf 45 Scanner by contacting a local digital photography distributor.
} } } } Does anyone have experiences (good or bad) with the Polaroid SprintScan 45? } } } } We want to use the scanner for getting high quality resolution scans from } } our sheet film TEM micrographs. } } } } Thanks in advance f snip, snip
A new option which I have not seen discussed on this forum is a drum scanner. The company ScanView which has made high-quality drum scanners for the pre-press market has now begun to market cheaper drum scanners at a relatively low price. We recently purchased the Scanview 3000 (3000 dpi, ~$15,000) which has an optical density range of 3.6, color resolution of 3 x 12 bit, 4096 levels, and a scanning area of 8.5" x 11.5". Some advantages of this scanner over the Leaf 45 are that the 3000 dpi resolution is available over the entire drum and the scan speed is much faster at the same resolution since it is a single pass scan. So, enlargements of an area of the negative which is off center are easy to do. You can also mount up to 6 negatives at once and set up batch jobs which will free up some of your time. We have been very pleased with the results so far. I believe it will deliver all possible information from the negative since the resolution of negatives are not much better than 3000 dpi in most circumstances. ScanView also makes two cheaper drum scanners the Scanmate Plus II (2600 dpi, o.d.=3.6) and the Scanmate Magic (2000 dpi, o.d.=3.0). I suggest that you contact your local digital photography dealer for a demonstration.
I have no financial interest in ScanView. I am just a satisfied user.
******************************************************* David F. Teter Los Alamos National Laboratory Materials Science and Technology: Metallurgy (MST-6) Mail Stop: G755 Los Alamos, NM 87545 ph: (505)665-0160 fax: (505) 665-0657 e-mail: teter-at-lanl.gov *******************************************************
I would like to ask several questions and I would be pleased if someone can answer one of them.
I want to capture images with an infrared film and capture only emitted wavelengths.
1) What literature can I read something about infrared emission wavelengths ?
2) What filters are available to select only a certain range of infrared wavelengths ?
3) Does anybody know any application of infrared photography in scientific work ?
Thank you for your attention.
Rui
/-------------------------------------------------------------------------\ | Rui Costa | e-mail: ruicosta-at-esb.ucp.pt | | Escola Superior de Biotecnologia |telephone: 351-2-5580044 | | Rua Dr Antonio Bernardino de Almeida |fax: 351-2-590351 | | 4200 PORTO | | | PORTUGAL | | \-------------------------------------------------------------------------/
Here's a can of worms if there ever was one! I'll infer that the reasons for the inquiry are that a) someone, probably your boss, has told you that you are not doing enough and b) you are already working as hard/fast as you can. Rate (call it turn around time or whatever) is only ONE measure of productivity. If it is the ONLY measure of productivity then your manager is failing to do his/her job properly. Turnaround time has to be evaluated along with quality, continuous improvement, customer satisfaction, profit/loss, employee morale, customer value, and a host of other criteria that are all inter-related. The analysis does not occur in a vacuum (bad pun) isolated from the rest of the world. You and your work are part of a system. Changing ANY part of that system WILL disrupt another part.
There has to be a consensus among ALL stakeholders as to what is an acceptable "workload." This consensus must be reached using FACT, not speculation, which can be obtained by benchmarking your lab against equivalent labs, then comparing best/worst cases. Of course the irony is that you be taking time away from the scope to look at best practices, etc. By including all stakeholders in a rational, calculated, cross-functional, balanced approach, everyone will have a better understanding of what can/can't be done and why/why not.
I know this answer does not give you a number and that is what you were looking for, but without a balanced approach including all stakeholders using hard data as well as personal input, you be in an increasingly tense loop leading to major problems.
Read anything by W. Edwards Deming, Peter Senge, Chris Argyris, Peter Drucker, Margaret Wheatley (sp?), Philip Crosby, Tom Peters....
IF YOU SPECIALIZE IN IMMUNOCYTOCHEMISTRY/IMMUNOHISTOCHEMISTRY/OTHER RELATED AFFININITY METHODS, may I URGENTLY draw your attention to my PROPOSED NEW NEWSGROUP to be called "sci.bio.immunocytochem"
The formal proposal can be found posted in the USENET NEWSGROUPS "news.announce.newgroups" and "news.groups". The latter contains material of a rather varied nature (!) but it is necessary to use this unmoderated group to hold set-up discussions for proposed new groups.
If you are keen to see A NEW IMMUNOCYTOCHEMISTRY NEWSGROUP, then please go to "news.groups", ignore all the rubbish, look carefully for the articles with "sci.bio.immunocytochem" in their subject line. By the time you read this message, the CALL FOR VOTES may well be underway. When you see "CFV: sci.bio.immunocytochem" (but not before please) then it is time to VOTE! You won't need access to Usenet to vote for my new group, only e-mail. We need at least 100 VOTES to get the group up and running!
For those people who are unfamiliar with Usenet, Newsgroups are a bit different from the Web or E-Mail. It is necessary to have access to "Usenet" which not all academic computer departments offer. So first check it you have access. Next you will need "news-reading software" which is available free by downloading it from the Web (or ask your computer department if they have some on a floppy disc). Once you have your "newsreader" (eg Free Agent for Windows) then you can read and post to newsgroups easily.
It is possible to access news via the Web, for example with Deja-news, but it is realy clunky and a last resort in my opinion. However, you CAN read the official proposal at the Royal Microscope Society web site {http://www.rms.org.uk} , or at the Introduction to Immunocytochemistry (Center for Cell Imaging Department of Cell Biology Yale University School of Medicine) web site {http://info.med.yale.edu/cellimg/CCIimmuno.html}
Proponent: Amanda Wilson {awilson-at-aw.u-net.com}
I am working with distarch phosphates (carbohydrate chains covalently linked together by Phosphorous), created with POCl3. I was hoping that someone can tell me whether it would be possible to use some type of probe (maybe with fluorescence?) to be able to detect the phosphorous based crosslinks. I have used SEM with EDS and this technique is not sensitive enough to detect the low level of phosphorous in the starch.
The oil drops on your EDS detector could be from either the diffusion pump, or from the rotary vane backing pump, or from both, and I don't know any easy way to tell which. If you can collect a few drops of it you might try putting it on a specimen stub and running an EDS analysis on it. If you find silicon to be present then you know that at least some of it comes from the diffusion pump. Even better, perhaps you can find someone in some chemistry lab that will run an infra red spectrem of it for you. That should give a difinitive identification.
In the past, we have had trouble with oil collecting on the EDS detector in one of our SEMs. and an FTIR spectral analysis showed it to be mainly from the rotary vane backing pump. Ed Birko, of the Hitachi service organization, has told me that on several occasions he has also found that the roughing pump was the major source of such contamination. As discussed on p. 144 of my book, "Vacuum Methods in Electron Microscopy," the molecules of the roughing pump oil get broken down into volatile low molecular weight fragments by the mechanical rubbing of the parts inside the pump, and these fragments readily migrate into the backing line and thence through the diffusion pump and into the vacuum system. Most diffusion pump oils, and particularly the silicone oils, are quite resistant to thermal and oxidative degradation, and so are unlikely to be a major contributor unless the vacuum system on your instrument is poorly designed or your diffusion pump has been subjected to one of the improper operating situations described on p. 214-223 of VM in EM.
There are two principal ways of dealing with contamination problems arising from the roughing pump. You can give your instrument a good cleaning and then install a good foreline trap, which must then be faithfully maintained to preserve its effectiveness (VM/EM p.147-149). [One approach that appears reasonably effective is to use the Micromaze traps sold by the Lesker Co,, heating them continuously with the heaters normally used for regeneration running at about 75% full power (VM/EM p. 147)] Alternatively, you can use an inert gas purging system, which is brought into operation when the instrument can be put into the standby mode, to attempt to sweep the oil molecules back out of the vacuum system - VM/EM p. 145 (XEI sell a system designed to perform this operation automatically).
In any event, you will need to remove the presently contaminating oil from your EDS detector window, otherwise it will begin to alter your detector sensitivity. This can be a very tricky process, and should be done with a great deal of care. It is best to check with the manufacturer of your detector for their recommendations before you try anything. (We have successfully removed oil from detectors with the standard beryllium windows by VERY CAREFULLY running a few drops of petroleum ether down over the window. Do NOT use a solvent such as acetone, or an alcohol, which may attack the epoxy normally used to hold the window in place, and be very careful not to puncture the window.)
Good luck,
Wilbur C. Bigelow, Prof. Emeritus Materials Sci. & Engr., University of Michigan Ann Arbor, MI 48109-2136 e-mail: bigelow-at-umich.edu; Fx:313-763-4788; Ph:313-764-3321
I agree with Tom Phillips-perhaps our suppliers could add some comments for us about age of these materials and possible smaller quantities available. It's been a long standing problem. Grace (Hello Stacie, Ted, Steve.....?)
Dear Folks, Please accept this message in hopes that it will answer most of your questions posted to me in the past two weeks. The real problem with Pb staining for epoxy sections is that almost nothing is known about the mechanism of heavy metal deposition on epoxides for TEM. We must first consider the interaction between the very active monomers in the section with the Pbcit. About 10% of all monomers stay unbound during a typical embedment. These monomers react with a variety of heavy metals - I have never had the time to prove that they will react detrimentally(for TEM) with Pbcit. However, I have repeatedly witnessed the situation where lead stain which did fine with random sections, repeatedly, consistently "dumped" when used on severely underpolymerized sections. Depending on the typed of tissue embedded and the care with which it was processed, much more than 10% of monomers remain unpolymerized. (This can be a huge problem when staining thick sections with the methylene or toluidine blues, since the monomers will engage the chromogens in undesirable redox shifts, sometimes resulting in rapid fading). So - before we even get to the lead, we have a vacillating situation. We must also note that not all Shell Epon 812 substitues are true substitues. Some replacements contain dilutents. This will affect the final staining nature of the sections, perhaps positively, perhaps negatively. We just do not know.
The pH of the lead solution is critical. Over four years of accurate record keeping, I was able to gradually intensify the Pb stain by dropping the pH. Finally I had nothing but precipitate. By increasing the pH of the lead stain, I could eventually eliminate all staining activity from the control sections.
One must control the pH. How this is done is immaterial. Very close control may be difficult when pellets are used - they are difficult to weigh, they absorb water. If one can use pellets and then titrate to the same pH every time, one will get reliably good results. If the relationship with pH and lead citrate is not understood, and pellets are used, a laboratory may experience variations in Pbcit staining intensity. (In 1983 I received over 400 requests for reprints for a simple paper I wrote regarding the use use of reliable staining practice. Over the years I have suggested that laboratories who turned to me with their propblems to do two things immediately: 1. Abondon pellets in favor or commercially titrated NaOH 2. Chealate the citrate and lead adequately by shaking and inversion for at least 25 minutes. This has solved most problems immediately.) If one uses commercially titrated NaOH, the worry about pH variations disappears.
It is thought that stored NaOH absorbs carbon dioxide from the atmosphere resulting in lead carbonate precipitate. Probably happens. However, I simply have not had any problems with it. Moreover, once when I accidentally bubbled about 10ml of air through about 5ml of Pbcit solution in a staining dish containing valuable sections, I expected the worst. Nothing happened. We breathe on our sections sitting in lead drops, etc. We do not boil water. It may be possible that if the Pb is at the correct pH and also the Pb is well chealated to the citrate (with adequate shaking and inversion),this problem is minimized. We do not worry about carbonate.
Well chealated lead citrate does not keep well in glass containers. We keep our lead solution in syringes in the refrigerator for 6 months. At this time I am using lead stain which I made up 18 months ago. I am still using it successfully. I do not recommed this, however. I am just very curious to how long it takes for the system to fall apart.
The original paper by Reynolds is wonderful storehouse of information. I recommend to everyone who is having any problem to get it and study it. Tomorrow I am sending out the chapter to those who requested it. If you did not get it in about 10 days (allow for snail mail), let me know.
If someone has unsolvable problems, please feel free to call me. I might be able to pinpoint something which is not obvious, since I (and my students) have made about every mistake possible with this lead soup. My phone # is 303-871-3026.
Bye. See you all in Cleveland at the MSA meeting, I hope! Hildy
H. Crossman wrote: } } Here's a can of worms if there ever was one! I'll infer that the } reasons for the inquiry are that a) someone, probably your boss, has } told you that you are not doing enough and b) you are already working as } hard/fast as you can. Rate (call it turn around time or whatever) is } only ONE measure of productivity. If it is the ONLY measure of } productivity then your manager is failing to do his/her job properly. } Turnaround time has to be evaluated along with quality, continuous } improvement, customer satisfaction, profit/loss, employee morale, } customer value, and a host of other criteria that are all inter-related. } The analysis does not occur in a vacuum (bad pun) isolated from the } rest of the world. You and your work are part of a system. Changing } ANY part of that system WILL disrupt another part. ...
I agree! Of course all of this depends on the environment you are in and what is expected from the results. One reason I do TEM in a research environment is that I am better at quality than quantity.
A few years ago when I was still very new to EM, I learned my lesson. I had started feeling bad about being a tortoise instead of a hare. I kept reading about all the quick processing and embedding techniques that are used in clinical labs, and thought "wow, I could save days of work!". Wrong. All I did was waste a lot of time and mess up a few months' worth of samples. My necessarily large blocks of tendon and ligament just didn't behave like small biopsy specimens. It turned out when I skimped on time, I got poor fixation, infiltration, staining, etc. and ended up with too many artifacts to do the image analysis necessary for my research project. I had to repeat many of the samples. So I don't mind being a tortoise any more; it might just save time in the long run. (Then there are the additional benefits of hiding in one's shell - read darkroom - when the boss comes by! If I could just get over my fear of can openers ... but that's another matter ...)
-Karen
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Life Sciences Sector Lab Reply: kszaruba-at-mmm.com 3M Company 3M Center 270-1S-01 Phone: 612-737-2971 St. Paul, MN 55144-1000 Fax: 736-1519
These opinions are my own and may not represent those of 3M.
We have a NORAN (Tracor Northern) 5400 Series II EDS system (Model Number TN-5402/BBAA) acquired in 1987. It does not have a hard drive and operates off of two floppy drives. It requires 5.25" double sided, high density floppy disks, the originals of which are no longer readable. They have not been in use other than to be the source of copies made periodically but apparently they have simply deteriorated with time. We have copies of copies that are working pretty well but I'm becoming concerned that they may crash/die and we'll have no good source from which to make new copies. NORAN does not have replacement masters of this vintage so I'm hoping to find someone who has original master disks in good working order who would be willing to loan them to be copied.
We need two disks of the set from the November, 1987 Release 1C. What will work with our system is very specific and the original label on the master disks reads:
SERIES II SOFTWARE RELEASE 1C NOV., 1987, TRACOR NORTHERN, INC.
Of the disks in this release, we need:
SQ/SSQ/PRZ/ZAF Master NOT BOOTABLE
and
MSCAN2 Master
Thanks in advance for any help or suggestions you may have.
___________________________________________________________________________ Barbara Reine, Botany Dept. Box 351330 Univ. of Washington, Seattle, WA 98195-1330 e-mail: reine-at-u.washington.edu; ph: (206) 543-1955 ____________________________________________________________________________
Does anyone out there know the origins of the word "Wehnelt". I have seen it stated in print that it is named after its inventor, though this individual was not identified. I have also seen it stated that it is simply the German word for "grid". I can confirm neither of these statements. Does anyone out there know for sure? If it was named after the inventor, who was this person? -- Fred Schamber ....................... mailto:fhscham-at-SGI.NET
I am trying to find a source of plastic pages for three-ringed binders which have pockets large enough to hold polaroid prints (these are 131mm x 106mm). Ideally each page should have 4 pocket and be able to fit two prints back-to-back so images are visible from both sides of the page.
I would really appreciate it if someone could give me an address/phone/fax/email of a supplier of these items (if, indeed, these things are made). Cheers,
Mark Blackford TEM Group Materials Division, ANSTO PMB 1, Menai, N.S.W. Australia 2234 Phone 61 2 9717 3027 Fax 61 2 9543 7179
Disclaimer: The views expressed in this E-mail message do not necessarily represent the official views of ANSTO from which this message was conveyed.
} Does anyone out there know the origins of the word "Wehnelt". I have } seen it stated in print that it is named after its inventor, though this } individual was not identified. I have also seen it stated that it is } simply the German word for "grid".
My 2 cents on this is that "Wehnelt" most definitely does not mean "grid" in German. It has no uses in German whatsoever beside "Wehnelt-Anode" in electron microscopy. (The word for "grid" is "Raster", as in "Rasterelektronenmikroskop", which is abbreviated REM--hence the Germans' tendency to write REM when they mean SEM in English.) Although I don't know anything about the inventor, I always assumed that the Wehnelt anode was named after it's inventor. It sure sounds like it could be a German last name, albeit not a very common one.
We are about to start a program of work, using the TEM, on the respiratory tract of mammals/humans and I was wondering whether anybody out there could recommend any references on the same (especially for healthy but also for diseased tissue).
We already have copies of Rhodin, A.G. (Histology a text and atlas) but I was wondering if anyone knew of text books/atlases or review papers, on ultrastructure, which were more specific. Our general searches have produced little so I thought that the readership, with its combined experience of a small planet, may be able to help or suggest new areas to search.
If you think that your reply will not be of general interest, please reply to me directly by e-mail at the address below.
thanks
Malcolm Haswell University of Sunderland UK e-mail: es0mhs-at-environment.sunderland.ac.uk (NB. 3rd character is zero)
I bought my sleeves from a local camera supply house. Can your Polaroid representative help? The product I use is called:
NegaFile P.O. Box 78 Furlong, Pennsylvania, USA 18925
Reorder number (part number?) 4504
The sleeves hold four Polaroid prints. One caveat, make sure the prints are dry or they will stick to the plastic.
------------------------------------------------ Opinions or statements expressed herein, rational or otherwise, do not necessarily reflect those of my employer.
Harold J. Crossman OSRAM SYLVANIA INC. Lighting Research Center 71 Cherry Hill Dr. Beverly, MA 01915 Phone: (508) 750-1717 E-mail: crossman-at-osi.sylvania.com
Our web sites: www.sylvania.com www.siemens.com --
I would like to know the experimental details of the gamma(electron) irradiation technique that has been used to enhance contrast in PE sections. Does anyone have this information ?
For the past two or three years, I have been using a text in my ultrastructure class entitled "Cell and Tissue Ultrastructure - A Functional Perspective," by P. C. Cross and K. L Mercer. It is a system by system atlas of ultrastructure. The chapter on the respiratory system has EM's on the mucosa, bronchioles, alveoli, type I and type II alveolar cells, components of the blood-air barrier, and alveolar macrophages. I have recommended the text to several of my colleagues throughout the state and they are now also using it. It has a 1993 copyright (ISBN 0-7167-7033-4) and is published by W. H. Freeman and Company. For the record, I have no interest, financial or otherwise, in W. H. Freeman or the authors of the text.
} We are about to start a program of work, using the TEM, on the respiratory } tract of mammals/humans and I was wondering whether anybody out there could } recommend any references on the same (especially for healthy but also for } diseased tissue). } } We already have copies of Rhodin, A.G. (Histology a text and atlas) but I } was wondering if anyone knew of text books/atlases or review papers, on } ultrastructure, which were more specific. Our general searches have } produced little so I thought that the readership, with its combined } experience of a small planet, may be able to help or suggest new areas to } search. } } If you think that your reply will not be of general interest, please reply } to me directly by e-mail at the address below. } } thanks } } Malcolm Haswell } University of Sunderland } UK } e-mail: es0mhs-at-environment.sunderland.ac.uk (NB. 3rd character is } zero)
Martin A. Levin Department of Biology Eastern Connecticut State University Willimantic, CT 06226 Phone: (860)465-4324 FAX: (860)465-5213
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Dear All,
I am trying to find a source of plastic pages for three-ringed binders which have pockets large enough to hold polaroid prints (these are 131mm x 106mm). Ideally each page should have 4 pocket and be able to fit two prints back-to-back so images are visible from both sides of the page.
I would really appreciate it if someone could give me an address/phone/fax/email of a supplier of these items (if, indeed, these things are made). Cheers,
Mark Blackford TEM Group Materials Division, ANSTO PMB 1, Menai, N.S.W. Australia 2234 Phone 61 2 9717 3027 Fax 61 2 9543 7179
Disclaimer: The views expressed in this E-mail message do not necessarily represent the official views of ANSTO from which this message was conveyed.
Mark,
We purchase polypropylene pages in serveral sizes to accomodate TEM negatives as well as SEM prints from 20th Century Plastics, P.O. Box 2376, Brea, CA 92622-2376, (800)767-0778. The following is a list of what we get: Item # EZV45-00 , EZ2C Archival holds 4 - 4x5" prints Item # EZVH4K-00, EZ2C Archival holds 6 - 4x6" prints Item # EZ116K-00, EZ2C Archival holds 12 - 3.5x4.5" negatives or prints
I hope this helps you. You can give them a call and they will send out a catalog of their products.
Pamela F. Lloyd Materials Science Engineer UES, Inc. 4401 Dayton-Xenia Rd. Dayton, OH 45432 (937) 255-1329 (Phone) e-mail: lloydpf-at-ml.wpafb.af.mil
Disclaimer: The views expressed in this e-mail message do not necessarily represent the official views of UES, Inc., and I have no connection to 20th Century Plastics other than being a user of their products.
Most of the original EM of lung (and most other organs) was done in the '60s and early '70s. Much of this material is too old to be in computer databases. Try the encyclopaedic Histology texts, Weiss for example, or Bloom and Fawcett. Also try the Handbook of Physiology; before embarking on the physiology of an organ they usually have a good chapter on LM and EM of it. Also try a big comprehensive textbook of pulmonary medicine, such texts often have an overview of LM and EM with references.
Geoff -- *************************************************************** Geoff McAuliffe, Ph.D. Neuroscience and Cell Biology Robert Wood Johnson Medical School 675 Hoes Lane Piscataway, NJ 08854 voice: (908)-235-4583; fax -4029 e-mail: mcauliff-at-umdnj.edu ***************************************************************
I may be dragging the thread away from contamination, but I thought that most electron microscope users avoided silicon based oils in diff pumps etc because they are extremely difficult to remove from the interior of an electron microscope and any contamination would normally have electrical insulating properties (catastrophic in an e.m.). Perhaps I am wrong but I would welcome any comments. After all this is one of the reasons why Santovac oils and their relatives became so popular (despite their costs).
If I am labouring under a mis-apprehension then I apologise.
Malcolm Haswell University of Sunderland UK ----------
The oil drops on your EDS detector could be from either the diffusion pump, or from the rotary vane backing pump, or from both, and I don't know any easy way to tell which. If you can collect a few drops of it you might try putting it on a specimen stub and running an EDS analysis on it. If you find silicon to be present then you know that at least some of it comes from the diffusion pump. Even better, perhaps you can find someone in some chemistry lab that will run an infra red spectrem of it for you. That should give a difinitive identification.
In the past, we have had trouble with oil collecting on the EDS detector in one of our SEMs. and an FTIR spectral analysis showed it to be mainly from the rotary vane backing pump. Ed Birko, of the Hitachi service organization, has told me that on several occasions he has also found that the roughing pump was the major source of such contamination. As discussed on p. 144 of my book, "Vacuum Methods in Electron Microscopy," the molecules of the roughing pump oil get broken down into volatile low molecular weight fragments by the mechanical rubbing of the parts inside the pump, and these fragments readily migrate into the backing line and thence through the diffusion pump and into the vacuum system. Most diffusion pump oils, and particularly the silicone oils, are quite resistant to thermal and oxidative degradation, and so are unlikely to be a major contributor unless the vacuum system on your instrument is poorly designed or your diffusion pump has been subjected to one of the improper operating situations described on p. 214-223 of VM in EM.
{snip}
Good luck,
Wilbur C. Bigelow, Prof. Emeritus Materials Sci. & Engr., University of Michigan Ann Arbor, MI 48109-2136 e-mail: bigelow-at-umich.edu; Fx:313-763-4788; Ph:313-764-3321
Silvia & listreaders: In an oil pumped system some oil will always find its way into the chamber but you need to minimise this. Recent contributions dealt with rotary pump trapping. Condensation will be severe if your EDS has a pinhole in the Be. Then, while the SEM is at atmospheric pressure, the EDS snout becomes quite cold and when the SEM is pumped any oil will condense "extra well". Your vacuum system cooling may not be the best either. A liquid N2 trap above the dif pump would prevent the problem but that is not a simple solution. Is your cooling water flowing well (checked at the outflow) and what is its temperature? I would prefer the outlet water to be no higher than 20 degrees and if you have a recirculating system set the temperature to 14-16 degrees. Dif pump heaters are generally designed for those temperatures. The other bad news is your oil. Silicone is wonderful in coaters, unfortunately it react with the e- beam and produces totally insoluble products and they result in uncleanable apertures and other column problems. That is why the world switched to other fluids about 25 years ago. Most used is Santovac5 - even though these fluids are not as tolerant of abuse, (poor vacuum when pump is hot= no, no) nor are they as cheap as the silicones. If you do not know how to clean the Be window, send me an email. Best of luck. Jim Darley
ProSciTech Microscopy PLUS PO Box 111, Thuringowa QLD 4817 Australia Phone +61 77 740 370 Fax: +61 77 892 313 Great microscopy catalogue, 350+ Links, MSDS ************************ http://www.proscitech.com.au } } Hi everyone, } } } I have two questions about type of oil you use for mechanical } and diffusion pumps for the microscopes. } } (I) We have a JEOL SM 35-C (SEM) since many years ago. In the } last two years we have often seen oil drops condensed on the EDS detector, } inside the chamber (at least it looks like oil, light green-dark yellow } coloured drops). } We've been using Dow Corning 704 silicon oil for the } diffusion pump since ever. Now, we're not sure what it is going on, we suspect } either: } * backstreaming problems from the mechanical pumps oil } * not sufficiently high vapor pressure for the DC 704 } } What do you think about it? any experience? } } } (II) We have been using Edwards 15 oil for the mechanical pumps. } This time we have gotten a very high quotation (according to our reduced budget) } for this oil. We're thinking of looking for other brand of the same type to } replace it. } Any idea what to buy? } } } We would appreciate to hear any suggestion from any of you. } } } Many thanks in advance. Sincerely, } } } } Silvia Montoro } Centro Regional de Investigacion y Desarrollo } Guemes 3450 } Santa Fe - Argentina } csedax-at-arcride.edu.ar
Steven and listreaders: I would not dare to argue with the great John Luft - but sometime back in the times when Watson "invented" and Reynolds and others perfected lead stain, (and remember that UA only was good enough for methacrylates but not the then new epoxies) I shared my lead staining problems with an inorganic chemist. I learned that dilute NaOH solutions and NaOH pellets absorb CO2; which on pellets can show as a powdery white layer. Ever since I have used the top layer of pellets from a jar for other purposes and used fresh pellets in boiled (now I would sonicate) H2O. I also learned that 10N NaOH (which is 40%) is happy without CO2 and DOES NOT ABSORB it. I think that I only made three batches of 10N NaOH - after moving to another lab. Stored in a solid plastic bottle that stuff is good for "1000 years". A one plus nine solution just before use provides fresh, near CO2 free 1 N sod. hydroxide. I never measured pH of the final Reynolds solution. My lead preparations always worked contamination free until exhausted - about a year.
Similarly, the suggested elaborate process to recrystallise UA is baffling. For decades I used about 5ml of saturated UA in a dropper bottle with a single drop of HCl added and stored in the fridge for a maximum of one month. That preparation too was intrinsically trouble free.
All experienced biomed TEM workers have had Pb and UA problems from time to time but mostly, the reasons are simple failures of normal techniques. These failures can be perplexing at times but I do not believe that we need to re-invent Pb or UA methods. Regards Jim Darley
ProSciTech Microscopy PLUS PO Box 111, Thuringowa QLD 4817 Australia Phone +61 77 740 370 Fax: +61 77 892 313 Great microscopy catalogue, 350+ Links, MSDS ************************ http://www.proscitech.com.au
one of the problems associated with lead citrate stain preparation is the presence of carbonate in the NaOH. John Luft came up with a solution for this years ago. Here is a copy of a handout from his 1976 seminar series that covers both uranyl acetate and lead citrate stain preparation.
......snip The source of the 1.0N Na0H is also a problem, since C02 adds rapidly to pellets of Na0H or to a stock solution. (10 N Na0H is available "carbonate-free" in polyethylene bottles. It may have been carbonate-free when it was made, but there is plenty of carbonate in it when it arrives). A neat device for ready access to carbonate-free Na0H at any time is the following:
Take a clean 250 ml polyethylene bottle and fill it about half full of Na0H pellets (from a freshly opened bottle, but this is not essential). Then fill it about 3/4 full with distilled water. Quickly cap and submerge in cold running tap water because much heat is generated. ......snip
I agree with Karen: a research environment is alot different than a clinical or quick-turn around lab. Research labs more often than not work with novel samples which require often novel approaches and slow, tedious processing as opposed to overnight processing of standardized tissue biopsies. Unfortunately, with major cutbacks for support of education and research, layers of accountants and administrators with no understanding of what goes on in these types of facilitities are having a larger say. I know it is our job to convince them that we cannot run identical to clinical or some commerical labs but it is getting extremely difficult to do so as when they only consider the bottom line ($$$). Of course this is not to say we shouldn't adopt ways to speed certain tasks in the lab, but these are usually in data base management and clerical duties. Getting back to the original question as far as work load, it depends on the lab type and particular study. I was once worked next to a histopath lab where 3 technicians cut paraffin blocks at about one per minute while another tech stained them (H&E) so the pathologists could read them by noon. This does not happen in a EM facility processing plant, fungal, spores, arthropods, embryos, assorted animal tissue, etc. for immunocytochemistry, autoradiography, image analysis, etc. and spending hours of beam time. Just my two cents Hank Adams EML New Mexico State University.
Hello to all, Many thanks to those who responded to my TEM stage drift inquiry.=20 Several of you requested a summary of the responses so here goes....there were ~10 responses from users & applications engineers.=20 The common denominator with respect to lack of stage stability was thermal stability as influenced by water temp. & air currents. =20 As to what levels of stability should be I expected from this instrument? The consensus is that within 10 - 30 min. the drift should be 1nm/min. or less & not more than a few nm/min. before that.=20 My compliments to those employed by vendors who replied. They were informative & I didn=92t suffer from salesman dispensing airware. =20
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } Hi } } I would like to ask several questions and I would be pleased if someone } can answer one of them. } } I want to capture images with an infrared film and capture only emitted } wavelengths. } } 1) What literature can I read something about infrared emission } wavelengths ? } } 2) What filters are available to select only a certain range of infrared } wavelengths ? } } 3) Does anybody know any application of infrared photography in } scientific work ? } } Thank you for your attention. } } Rui } } /-------------------------------------------------------------------------\ } | Rui Costa | e-mail: ruicosta-at-esb.ucp.pt | } | Escola Superior de Biotecnologia |telephone: 351-2-5580044 | } | Rua Dr Antonio Bernardino de Almeida |fax: 351-2-590351 | } | 4200 PORTO | | } | PORTUGAL | | } \-------------------------------------------------------------------------/ } } } Hello Rui:
The easiest publications to start with are the Kodak series,N-17 "Infrared Films", M-28 "Applied Infrared Photography, N-1 "Medical Infrared Photography" and B-3 "Kodak filters for Scientific and technical use". You can get them through photography stores.
89B is the visibly opaque filter you would use to record just infrared. Since infrared film will record wavelengths in the visible spectrum as well as in the range of 700-900 nm.
We use infrared in dermatology to record the patterns of blood vessels in the skin. This is possible because the longer wavelengths can penetrate deeper.
Bob Underwood Morphology Core U of Washington Seattle
We buy our plastic storage pages from the University bookstore and also from local photography stores. They are archival quality and come in a variety of sizes. They are made by Print File Archival Preservers, PO Box 607638, Orlando Fl, Ph:407-886-3100; Fax 407-886-0008. The type I use for 4x5 polaroid prints is product #45-8P (which are a heavier weight than the negative preservers).
Hope this helps.
Rosemarie Rosell Assistant Research Scientist Dept. of Plant Sciences University of Arizona Tucson,AZ 85721 Ph: 520-621-1230 Fax: 520-621-8839
On Wed, 30 Apr 1997, Mark Blackford wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Dear All, } } I am trying to find a source of plastic pages for three-ringed binders } which have pockets large enough to hold polaroid prints (these are 131mm x } 106mm). Ideally each page should have 4 pocket and be able to fit two } prints back-to-back so images are visible from both sides of the page. } } I would really appreciate it if someone could give me an } address/phone/fax/email of a supplier of these items (if, indeed, these } things are made). Cheers, } } Mark Blackford } TEM Group } Materials Division, ANSTO } PMB 1, } Menai, N.S.W. } Australia } 2234 } Phone 61 2 9717 3027 } Fax 61 2 9543 7179 } } Disclaimer: } The views expressed in this E-mail message do not necessarily represent the } official views of ANSTO from which this message was conveyed. } } }
} I am trying to find a source of plastic pages for three-ringed binders } which have pockets large enough to hold polaroid prints (these are 131mm x } 106mm). Ideally each page should have 4 pocket and be able to fit two } prints back-to-back so images are visible from both sides of the page. } I would really appreciate it if someone could give me an } address/phone/fax/email of a supplier of these items (if, indeed, these } things are made).
These things are called "Polyview Pages" and are available sized to hold 4 Polaroids or 6 3-1/4" x 4" negatives. They are available from us, and, I believe, from several other EM supply companies.
Best regards, Steven E. Slap, Vice-President ******************************** Energy Beam Sciences, Inc. Adding Brilliance To Your Vision ebs-at-ebsciences.com http://www.ebsciences.com/ ********************************
What are some effective solvents for removing adhesive tape, spots, etc. from aluminum SEM mounts. Acetone is marginal as are other ketones. I've tried several alcohols without success. Caustics are out because they eat the aluminum. Pure and mixed acids don't work either. I've even tried WD-40 and liquid dish soap.
The ideal solvent would have low toxicity, low cost, etc... Water solubility would be a bonus. I'd really like a chlorinated solvent vapor degreaser! Fat chance.
Any suggestions? Terpenes, perhaps?
------------------------------------------------ Opinions or statements expressed herein, rational or otherwise, do not necessarily reflect those of my employer.
Harold J. Crossman OSRAM SYLVANIA INC. Lighting Research Center 71 Cherry Hill Dr. Beverly, MA 01915 Phone: (508) 750-1717 E-mail: crossman-at-osi.sylvania.com
Our web sites: www.sylvania.com www.siemens.com --
YES, the do exist. We use a product called Kleer-Vu, Proline #14912 plastic, 3-ring binder pages for 4x5 prints. In fact, we use it for our Polaroid prints, back-to-back and it works beautifully. You should be able to purchase these at any professional graphic supplier. If you have problems finding a vendor, contact me again.
} I am trying to find a source of plastic pages for three-ringed binders } which have pockets large enough to hold polaroid prints (these are 131mm x } 106mm). Ideally each page should have 4 pocket and be able to fit two } prints back-to-back so images are visible from both sides of the page. } } I would really appreciate it if someone could give me an } address/phone/fax/email of a supplier of these items (if, indeed, these } things are made). Cheers, } } Mark Blackford } TEM Group } Materials Division, ANSTO } PMB 1, } Menai, N.S.W. } Australia
#################################################################### John J. Bozzola, Ph.D., Director Center for Electron Microscopy Neckers Building, Room 146 - B Wing Southern Illinois University Carbondale, IL 62901-4402 U.S.A. Phone: 618-453-3730 Fax: 618-453-2665 Email: bozzola-at-siu.edu Web: http://www.siu.edu/departments/shops/cem.html ####################################################################
} Does anyone out there know the origins of the word "Wehnelt". I have } seen it stated in print that it is named after its inventor, though this } individual was not identified. I have also seen it stated that it is } simply the German word for "grid". I can confirm neither of these } statements. Does anyone out there know for sure? If it was named after } the inventor, who was this person? } -- Dear Fred, About two months ago I replied to a Gustavo Waenelt about this. Here is my reply:
Harold: I have a product called Ease-Away, (manufactured by Wood Life Ltd, 9331 Park Lane, Franklin Park, IL 60131) which I bought from some mail order catalog a few years back whose advertising claims that it "removes adhesive and adhesive residue from any surface quickly, easily, and safely", and which I have, in fact, found to do a very good job on adhesive tape residues, but to work more slowly on removing tape itself (which is not surprising, since the tape represents a pretty heavy dose of plastic of some kind or the othger to dissolve).
Incidentally, I have also found that my pet cleaning agent, Tilex Soap Scum Remover, works pretty well on tape residues. As I've mentioned in previous comments on this listserver, it also does a pretty good job of removing silicone and polyphenyl ether diffusion pump oils from metal surfaces, paint from your hands, oil stains from carpets, food stains from clothing, etc.
Wilbur C. Bigelow, Prof. Emeritus Materials Sci. & Engr., University of Michigan Ann Arbor, MI 48109-2136 e-mail: bigelow-at-umich.edu; Fx:313-763-4788; Ph:313-764-3321
20th Century Plastics, 205 So. Puente St., Box 2393, Brea, CA 92622-2393, Tel: 1-800-767-0777; Fax: 1-800-786-7939, sell a wide variety of plastic sheets for jholding a wide variety of different size items. You might check with them to see if they have something to meet your needs. (They list stock No. V45000A-00 as having 4.5x5.25" pockets suitable for holding Polaroid prints.) Good luck,
Wilbur C. Bigelow, Prof. Emeritus Materials Sci. & Engr., University of Michigan Ann Arbor, MI 48109-2136 e-mail: bigelow-at-umich.edu; Fx:313-763-4788; Ph:313-764-3321
Its interesting that you mention this. I never look forward to having to clean the tape off of my aluminum stages. The Acetone is labor intensive and time consuming. Just this morning I put a few mounts that had several layers of carbon tape on them and placed them in a platinum dish and set them in a muffle furnace -at- 300 C. After about an hour the tape had charred, but looked as though it was still going to take some scraping to get off. I increased the temperature to 400 C and left it for another hour. The tape had powdered and fallen off of the stages. I rinsed them in DI water and set them in an ultrasonic bath and they came clean.
Have you tried the commerial product "Goo-begon"? Its generally available in most N.American harware stores, painting stores, and grocery stores as a product to removing "Sticky residues left behind after removing tapes and labels".
Richard E. Edelmann, Ph.D. Electron Microscopy Facility Supervisor 352 Pearson Hall Miami University, Oxford, OH 45056 Ph: 513-529-5712 Fax: 513-529-4243 E-mail: edelmare-at-muohio.edu
"WE ARE MICROSOFT. RESISTANCE IS FUTILE. YOU WILL BE ASSIMILATED."
R. Breeze and M. Turk; 1984; Cellular structure, function and organization in the lower respiratory tract; Environmental Health Perspectives 44:3-24, 1984.
J. A. Nowell and W.S. Tyler; 1971; Scanning electron microscopy of the surface morphology of mammalian lungs; Amer. Review of Respiratory Disease 103:313-328, 1971.
Also, look for more recent papers by C. Plopper, J. Crapo, or K. Pinkerton.
Jane A. Fagerland, Ph.D. Dept. Microscopy and Microanalysis Abbott Laboratories Abbott Park IL 60064
} What are some effective solvents for removing adhesive tape, spots, etc. } from aluminum SEM mounts. Acetone is marginal as are other ketones. } I've tried several alcohols without success. Caustics are out because } they eat the aluminum. Pure and mixed acids don't work either. I've } even tried WD-40 and liquid dish soap. } } The ideal solvent would have low toxicity, low cost, etc... Water } solubility would be a bonus. I'd really like a chlorinated solvent } vapor degreaser! Fat chance. } } Any suggestions? Terpenes, perhaps? } Dear Harold, I'm surprized that acetone and ethanol did not remove adhesive. The adhesive from tape in our lab is readily removed by these. I'd try an aromatic, preferrably xylene or toluene--used in a hood, of course. Avoid benzene--it's much more toxic than the hydroxylated aromatics. Freon is another possibility. I'd also avoid chlorinated solvents if possible because of toxicity. As you said, stay away from bases, including strong detergents. Fine abrasives are another option; something like wehnelt-polishing compound can be used. Good luck. Yours, Bill Tivol
} Does anyone know the dangers associated with a technique called Confocal } Microscopy or something like that? In our Chemistry Dept. we have a } research group that works with a Class 4 LASER. To the best of my } understanding, the LASER beam is directed toward a sample that is sitting } under a microscope. Some of the beam is reflected and filtered so the } power or strength of the beam is reduced before it hits the sample } (a biological sample). The researcher then looks through the microscope } objective at the sample. } } Currently, we have a LASER safety program in place. This particular } procedure concerns me because this technique allows the researcher to } look directly at the LASER beam. This does not sound too good. Also, } the microscope objective may magnify the beam and direct the beam } toward the researchers eye. } } Currently, the LASER that is being used is an Class 4 ARGON Ion LASER } (Continuous Wave) with a wavelength of 0.488 um (ultraviolet range) and a } beam power of 1.5 W. } } Does anyone know where I can get more information on this subject. I am } particularly interested in the safety aspects of this technique. It } may be the case that the LASER beam hitting the sample has a low enough beam } power to classify it as a Class 1 LASER. } } Thanks in advance for your help. } } Laurie Princiotto } Laboratory Safety Specialist } Indiana University - Dept. of Environmental Health and Safety } Phone: (812)-855-6115 } Fax: (812)-855-7906 } E-mail: lprincio-at-indiana.edu
Laurie, This does not sound like a very safe activity to me. I would be very concerned until proven safe.
Regarding the lasers used in confocal microscopes...I've worked with the Bio Rad commercial system and I believe most use a class II Ar or Ar/Kr laser at ~1mW power (457-648nm). The laser image cannot be directly observed but is displayed in a computer screen.
You're best info sources would be the manufacturers of confocal microscopes.
Edward J. Basgall, PhD The Pennsylvania State University Surface Chemistry Group ejb11-at-psu.edu Materials Research Institute Building Ph: 814-865-0493 University Park, PA 16802-7003 FAX: 814-863-0618
My guess is that the citrus-derived "limonene" product used as a substitute for xylene in histology should work fine. It solubilizes paraffin, oils, etc. and is very safe. Most histo labs use this solvent these days, so drop by you local histo lab and give it a try.
} What are some effective solvents for removing adhesive tape, spots, etc. } from aluminum SEM mounts. Acetone is marginal as are other ketones. } I've tried several alcohols without success. Caustics are out because } they eat the aluminum. Pure and mixed acids don't work either. I've } even tried WD-40 and liquid dish soap. } } The ideal solvent would have low toxicity, low cost, etc... Water } solubility would be a bonus. I'd really like a chlorinated solvent } vapor degreaser! Fat chance.
#################################################################### John J. Bozzola, Ph.D., Director Center for Electron Microscopy Neckers Building, Room 146 - B Wing Southern Illinois University Carbondale, IL 62901-4402 U.S.A. Phone: 618-453-3730 Fax: 618-453-2665 Email: bozzola-at-siu.edu Web: http://www.siu.edu/departments/shops/cem.html ####################################################################
Thanks to all who responded to my inquiry of March 19 concerning embedding and sectioning, I have summarized the responses below.
On the use of silane adhesion promoters (in my case Dow Corning Z-6040) for embedding in epoxy: Phil Swab uses these promoters on materials specimens. He uses 1% Z-6040 in 1:1 methanol:water for an hour then dries the specimen. He then embeds in epoxy with a replacement of half the amine polymerization initiator ( eg. DMP-30) with Z-6040. Stacie Kirsch of EMS, has used Z-6040 with epoxies and found no difference between conventional embedding and use of Z-6040. I ran an experiment using some bird feather barbs using EM-BED 612 (EMS) with the following protocols: In cases 1 & 2 acetone was the transitional solvent, and specimens were left in 1:1 acetone:epoxy overnight and in other acetone epoxy steps for a minimum of 4 hrs, and 4 hrs in the epoxy alone before transferring to the final epoxy embedment. 1. No Z-6040. 2. 1% Z-6040 added to all EtOH steps up through 100% EtOH but not in acetone. 3 Phil Swab method see above. To summarize, there were no differences in appearance under the beam. All blocks cut easily and equally well, and all suffered from the same problem. The problem was that on pickup on uncoated grids the feather sections fell out of the block. Mounting on coated grids was mandatory. If you use Z-6040, LRWhite appears to be the plastic of choice. If anyone has an experimental bent, trying some of the other adhesion promoting silanes may work. A long preembedding period in solvent+epoxy and in the pure epoxy certainly seems to help regardless of the other embedding details. Thanks to Caroline Schooley for suggesting Phil Swab.
Concerning cells on membranes: Neelima Shah suggested letting the membrane roll up (usually happens in higher alcohols or acetone in my experience) and vacuum infiltration. Wis Jablonski suggests using a hard Spurr mixture with long infiltration and the Spurr only step under mild vacuum. Karen Zaruba suggests the use of LRWhite and heat polymerization. Laura Patrone suggests cutting the block so that the cell layer is down (the first part of the specimen to be cut by the knife). Many of these correspondents gave suggestions for membrane inserts such as Millepore HA membranes, Millicel CM membranes. My problem was that I am restricted to the membranes being used by the investigators, and in the last batch I had, there was essentially no adhesion between the cell layer and the membrane, they split apart during the trimming (as soon as any shearing force was applied in the general vicinity). For the more tractable membranes the change in orientation of the block with respect to the knife seems a reasonable easy fix. Also longer infiltrations in plastic+ solvent and in plastic infiltration step work better. LRWhite is also worthy of consideration by the epoxyheads out there, such as myself.
I have no pecuniary connection to Dow Corning or EMS.
Bruce Cutler, Microscopy Laboratory, University of Kansas, Lawrence
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } What are some effective solvents for removing adhesive tape, spots, etc. } from aluminum SEM mounts. Acetone is marginal as are other ketones. } I've tried several alcohols without success. Caustics are out because } they eat the aluminum. Pure and mixed acids don't work either. I've } even tried WD-40 and liquid dish soap. } } The ideal solvent would have low toxicity, low cost, etc... Water } solubility would be a bonus. I'd really like a chlorinated solvent } vapor degreaser! Fat chance. } } Any suggestions? Terpenes, perhaps? } } } ------------------------------------------------ } Opinions or statements expressed herein, rational or otherwise, do not } necessarily reflect those of my employer. } } Harold J. Crossman } OSRAM SYLVANIA INC. } Lighting Research Center } 71 Cherry Hill Dr. } Beverly, MA 01915 } Phone: (508) 750-1717 } E-mail: crossman-at-osi.sylvania.com } } Our web sites: www.sylvania.com } www.siemens.com } -- } } "Crossman, Harold" {crossman-at-osi.SYLVANIA.com} } } "Crossman, Harold" {crossman-at-osi.SYLVANIA.com} } Harold, have you tried soaking them in a 10% solution of Microsolution? This has worked for me several times I've wanted to reuse stubs. Use hot water (the hotter the better) when starting then add the Microsolution. It works best with hot water but letting things soak in cold water overnight works O.K. too. Hope this helps.
Peace through Christ,
Phil Rutledge, Director e-mail: prutle1-at-gl.umbc.edu Center for Microscopy voice: (410) 455-3582 UMBC Dept. of Biology fax: (410) 455-3875 Catonsville, MD 21228 ///// / / / / /////// /////// / / /////// /////// / / / / / / / / / / / / /////
I may be dragging the thread away from contamination, but I thought that most electron microscope users avoided silicon based oils in diff pumps etc
because they are extremely difficult to remove from the interior of an electron microscope and any contamination would normally have electrical insulating properties (catastrophic in an e.m.). Perhaps I am wrong but I would welcome any comments. After all this is one of the reasons why Santovac oils and their relatives became so popular (despite their costs).
If I am labouring under a mis-apprehension then I apologise.
Malcolm Haswell University of Sunderland UK
All of the silicon based DP fluids I am aware of (which may not be all that are on the market) will break down under an electron beam and deposit a layer similar to glass on the nearest cool surface in the column. I don't know of any EM manufacturers that would recommend their use because they are almost impossible to remove if they do get in the column. We don't even use silicon based fluids in our vacuum evaporator.
Years ago I used to make carbonate-free NaOH by dilution of aliquots drawn from a saturated solution of NaOH pellets, the procedure came from the classic text on quantitative inorganic analysis by Vogel. The story is that Na2CO3 is more-or-less totally insoluble in saturated NaOH. I can't remember what concentration NaOH the saturated solution is, and, of course you have to dilute it with recently-boiled out water, but at least the problem of getting carbonate-free NaOH is thus eliminated.
cheers
Ritchie
Ritchie Sims phone: 64 9 3737599 ext 7713 Department of Geology fax: 64 9 3737435 University of Auckland Private Bag 92019 Auckland New Zealand
Malcolm Haswell wrote: =============================================== .............. but I thought that most electron microscope users avoided silicon based oils in diff pumps etc because they are extremely difficult to remove from the interior of an electron microscope and any contamination would normally have electrical insulating properties (catastrophic in an e. m.). Perhaps I am wrong but I would welcome any comments. After all this is one of the reasons why Santovac oils and their relatives became so popular (despite their costs).
If I am labouring under a mis-apprehension then I apologise. ================================================= If you are laboring (sorry labouring) under a "mis-apprehension" then you are not alone!
The only time I have ever thought that the Dow Corning silicone based fluids would be acceptable for use in an EM lab was in the vacuum evaporator, unless of course you were interested in doing Si analyses by EDS and you did not want there to be even remotely the possibility of having Si contamination from the pump fluid. There are some who argue for other reasons even against the vacuum evaporator use.
But I have heard far too many horror stories over the years of columns being flooded with silicone DP fluid and then having the microscope undergo a month long or more cleaning job. Stick with a polyphenyl ether fluid, such as Santovac (R) or if you are really on a super tight budget, use dioctylsebacate (e.g. Octoil-S (R)) which are of course easy to clean up in the event of a column catastrophe.
Chuck
=================================================== Charles A. Garber, Ph. D. Ph: 1-(610)-436-5400 President 1-(800)-2424-SPI SPI SUPPLIES FAX: 1-(610)-436-5755 PO BOX 656 e-mail: cgarber-at-2spi.com West Chester, PA 19381-0656 USA Cust. Service: spi2spi-at-2spi.com
Look for us! ############################ WWW: http://www.2spi.com ############################ ==================================================
One of my colleagues is looking for the excitation and emission spectra for fluorogold. The supplier gives out the Ex & Em peaks but says they don't have a full spectrum. Has anyone out there generated their own? TIA.
Thomas E. Phillips, Ph.D. Associate Professor of Biological Sciences Director, Molecular Cytology Core Facility 3 Tucker Hall University of Missouri Columbia, MO 65211 (573)-882-4712 (voice) (573)-882-0123 (fax)
We are preparing to purchase an ultramictrome for our centralized EM facility. This instrument will be used by a variety of people, including undergraduate students. If you have recently purchased such an instrument, we would appreciate your comments, good and bad. Pricing information would also be helpful.
Bill McManus Department of Biology Utah State University Logan UT 801 797 1920
Aloha! You can usually count on me to wade in with the High Humidity viewpoint...
We use Polaroid Type 55 positive/negative film, which requires the positives to be coated. In our usual high humidity environment the coating never really dries forever, and rehydrates easily. The coating on prints stored in various kinds of sleeves often ends up sticking to the material and, when the prints are peeled out, exhibits unsightly marks. I called Polaroid and got a list of acceptable materials that could come into contact with the coating and which included such things as ceramic and some plastics but not others. (Please call them rather than make me dredge the file cabinet for this list!).
I conducted a year-long experiment with prints stored in 3-ring storage sheets from various companies, made of various materials, left under stacks of heavy books. The sheets that work best under my conditions are Polyethylene Storage Sheets for 4X5 Negs and Prints, Order #4504, from
NegaFile P.O. Box 78 Furlong, PA 18925 215-348-2356
This is not to say that sheets from other sources won't be just fine for uncoated prints or coated prints in areas with lower humidity!
It's a beautiful, sunny day with a humidity of 70 percent IN THE LAB, and south shore surf is coming up.
Tina
http://www.pbrc.hawaii.edu/bemf/microangela **************************************************************************** * Tina (Weatherby) Carvalho * tina-at-pbrc.hawaii.edu * * Biological Electron Microscope Facility * (808) 956-6251 * * University of Hawaii at Manoa * http://www.pbrc.hawaii.edu/bemf* ****************************************************************************
the response to my recent question about polaroid print storage was overwhelming. Thanks to those who responded I now have several lines of enquiry, even a couple in Australia. Cheers,
Mark Blackford TEM Group Materials Division, ANSTO PMB 1, Menai, N.S.W. Australia 2234 Phone 61 2 9717 3027 Fax 61 2 9543 7179
Disclaimer: The views expressed in this E-mail message do not necessarily represent the official views of ANSTO from which this message was conveyed.
the response to my recent question about polaroid print storage was overwhelming. Thanks to those who responded I now have several lines of enquiry, even a couple in Australia. Cheers,
Mark Blackford TEM Group Materials Division, ANSTO PMB 1, Menai, N.S.W. Australia 2234 Phone 61 2 9717 3027 Fax 61 2 9543 7179
Disclaimer: The views expressed in this E-mail message do not necessarily represent the official views of ANSTO from which this message was conveyed.
There is another option for scanning film negatives. A Leaf MicroLumina camera can be coupled with a high frequency light box and a copy stand to provide a simple and highly flexible solution to the problem of scanning film and transparencies.
The camera can be racked up and down on the stand to provide a field of view from 1" X 1.5" up to 9" X 12". The camera has a resolution of 2700 X 3380 pixels and can capture 12 bit grayscale and 36 bit color.
This configuration is currently offered by our company to the electron microscopy community under the name TEMSCAN.
If anyone is interested in this product, please contact us by phone: 516-773-4305 or e-mail: sales-at-electroimage.com.
Thanks to the several people who responded to my question.
The following was received directly from Bart Cannon and has some useful detail. I thought others might be interested.
} Arthur Wehnelt (1871-1944) was the German physicist who invented the } "Hot Cathode" electron tube which employed what was called a "grid" to } direct a stream of electrons. It corresponds fundamentally to the } electron gun used in oscilloscopes, TVs, and electron microscopes. Grid } and wehnelt have been interchangable terms to some degree.
-- Fred Schamber ....................... mailto:fhscham-at-SGI.NET
If you have "copies of copies" which work ok, and you have suitable blank media, there is no reason you cannot copy these again to generate archive disks. If you have a "verify" command/switch - use it. ...Or am I missing something??
------------------------------------------------------------------------ The Microscopy ListServer -- Sponsor: The Microscopy Society of America
We have a NORAN (Tracor Northern) 5400 Series II EDS system (Model Number TN-5402/BBAA) acquired in 1987. It does not have a hard drive and operates off of two floppy drives. It requires 5.25" double sided, high density floppy disks, the originals of which are no longer readable. They have not been in use other than to be the source of copies made periodically but apparently they have simply deteriorated with time. We have copies of copies that are working pretty well but I'm becoming concerned that they may crash/die and we'll have no good source from which to make new copies. NORAN does not have replacement masters of this vintage so I'm hoping to find someone who has original master disks in good working order who would be willing to loan them to be copied.
We need two disks of the set from the November, 1987 Release 1C. What will work with our system is very specific and the original label on the master disks reads:
SERIES II SOFTWARE RELEASE 1C NOV., 1987, TRACOR NORTHERN, INC.
Of the disks in this release, we need:
SQ/SSQ/PRZ/ZAF Master NOT BOOTABLE
and
MSCAN2 Master
Thanks in advance for any help or suggestions you may have.
___________________________________________________________________________ Barbara Reine, Botany Dept. Box 351330 Univ. of Washington, Seattle, WA 98195-1330 e-mail: reine-at-u.washington.edu; ph: (206) 543-1955 ____________________________________________________________________________
While we're on the subject, and inspired by Chuck's comments about use of same in vacuum coaters, you may recall that about a year ago I asked about DC 704 in a coater used for carbon coating mineral samples for subsequent EDS analysis. I mounted up some graphite rod, analysed it
a uncoated b coated as usual ie with LN2 trap c as b, but left under diff pump vacuum for an extra hour d as c, but left under diff pump vacuum without LN2 in the trap for 2 hours(!). This was with a more-than-9-years-old charge of DC704 in the Edwards 306 coater.
Subsequent analysis showed no detectable Si in any of the samples.
So I cleaned out the pump, using Will Bigelow's favourite Tilex (thanks very much for the sample, Will) then Trichloroethylene, recharged it with DC704, and everything is hunky-dory.
In the diff pump of my JEOL JXA-5A, I use the JEOL-recommended Lion S oil, which I think is just a hydrocarbon oil, it's pretty cheap, and I get a vacuum better than 5 times 10 to the minus 6 torr. I change it every year.
cheers
Ritchie
Ritchie Sims phone: 64 9 3737599 ext 7713 Department of Geology fax: 64 9 3737435 University of Auckland Private Bag 92019 Auckland New Zealand
Check with the Manufacturer of your SEM or the maker of your ODP for the proper oil. Hopefully it did not come from the factory with the silicone stuff. The type and quantity of oil should match the Pump. Typically Sanovac requires a hotter (higher Wattage) heater than Octoil. If you put Octoil in with a hotter heater you may have charcoal. And if you use a cooler heater with Sanovac you may never pump down.
It seems to me that if you have major back-streaming that you will see some oil elsewhere and be cleaning your column often to maintain decent astigmatism. I would recommend checking the detector window and eliminating the Silicone oil, before you have more problems.
I am not a chemist, but... I use Skelly B (i.e., hexanes?) in our lab to remove adhesive. I particularly use it to clean up after using pressure-sensistive adhesive polishing papers on our aluminum wheel. It is one of the few things that cut the adhesive at all.
At 11:42 AM 4/30/97 -0400, you wrote: } What are some effective solvents for removing adhesive tape, spots, etc. } from aluminum SEM mounts. Acetone is marginal as are other ketones. } I've tried several alcohols without success. Caustics are out because } they eat the aluminum. Pure and mixed acids don't work either. I've } even tried WD-40 and liquid dish soap. } } The ideal solvent would have low toxicity, low cost, etc... Water } solubility would be a bonus. I'd really like a chlorinated solvent } vapor degreaser! Fat chance. } } Any suggestions? Terpenes, perhaps? ---------------------------------------------------- Warren E. Straszheim 270 Metals Development, Ames Lab/ISU, Ames IA, 50011 Phone: 515-294-8187 FAX: 515-294-3091
Try an oily solvent such as Histolene. Its extracted from citrus and has a pleasant smell and is reputed to be of very low toxicity. Its sold as a substitute for xylene which is none of the above. OR eucalyptus oil. These are also good for getting bandaids off small children with a minimum of weeping.
I am seeking some input and suggestions on a problem that I am having with our Denton Desk II cold sputter coater.
About 2 weeks ago, a student came to inform me that the sputter button wasn't "popping" when depressed as usual - I believe the popping is the opening of the solenoid valve which admits Argon to the chamber. After confirming that the problem existed, I noted that the outlet valve from the regulator had been turned off (I'm not sure if this contributed to the situation or not). Upon opening the back of the unit, I found a blown 2 Amp slo-blow fuse.
After replacing the fuse, the sputter valve has been working just fine, however, after replacing it, a new problem arose - when the 45mA current was applied to the gold target (at 50 millitorr), the typical plasma glow was not evident even though the indicator read 45mA - I did notice some initial arcing, then nothing!
After consulting a very helpful sales/service representative (who suggested that I take the sputterhead apart and clean the teflon spacer/insulator) I was able to restore normal function - at least it seemed to be fixed for a few trial runs!
Now, no matter how many times I take the head apart and clean each piece, the coater fails after only a few runs or less - it arcs violently and the solenoid valve automatically shuts down (cycles to off position).
Has anyone had similar problems with the unit and/or any advise? This unit was purchased in 1992 and is not used excessively (1 to 2 SEM courses per year) - I recently replaced only my second gold target in that amount of time.
Thanks in advance for your kind assistance.
Stephen J. Beck Associate Professor Bio-Imaging Center/Electron Microscopy Department of Biology Nassau Community College Garden City, NY 11530 Voice Mail: (516) 572-7829 Email: {becks-at-sunynassau.edu} URL: {http://www.sunynassau.edu/webpages/biology/becks.htm}
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