The Microscopy & Microanalysis '97 Search Engine is now on-line at
http://www.msa.microscopy.com
using this engine you may search the M&M' 97 program by Author, Title Keywords, Symposium Name or Day of the Week and find out when and where each presentation will occur.
Contrary to last year, the abstracts for all papers for the meeting will not be on line this year as the publication and organization of that aspect of the meeting is being coordinated by our new Journal.
A committee within the Dutch Microscopy Society (NVvM) is working on the general issue of calibration of microscopes (SEM, TEM, LM, AFM, STM). Special attention is paid to calibration in relation with accreditation. We would like to contact a microscopy lab that is certified or users of certified microscopes in order to exchange some ideas. Any general information on the issue is welcomed as well.
Thanks in advance.
Bart Nelissen Akzo Nobel Central Research Dept. Applied Physics Microscopy eMail: Bart.B.J.Nelissen-at-akzo.nl
I think it should work fine if both monoclonals are directly conjugated. But if you are using conjugated secondaries you will get your second primary sticking to your first secondary. You will also get your second secondary sticking to your first primary. So you will get various states of crossover. I think after you put on your first primary and secondary, block with 10x excess of anti-mouse Fab fragments to bind up all the free sites should work.
Bob Underwood Morphology Core U of W Seattle, WA, USA
On Fri, 30 May 1997, Geoff McAuliffe wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Dear Microscopists: } } Why is it that when I do double immunostaining with two } successive monoclonals I get inconsistant results with 'stains' that are } fine by themselves? If I do a polyclonal followed by a monoclonal, no } problems. What is the story? Are there ways around this? } Thanks in advance! } } 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 } *************************************************************** }
Steve, In the past I have purchased water filters and particle filters for CPD from Tousimis Res. Corp. 1-800-638-9558 is the last ph.# I have for them. (No affiliation, of course) cheers ed
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
} any suggestions where I can buy a water trap for my liquid carbon dioxide } tank attached to my CPD? } } TIA } } steve } } --------------------------------------------------------------------- } } Dr. Steven Barlow } EM Facility/Biology Department } 5500 Campanile Drive } San Diego CA 92182-4614 } phone: (619)594-4523 } fax: (619) 594-5676 } email: sbarlow-at-sunstroke.sdsu.edu } website: http://www.sci.sdsu.edu/EM_Facility
Please can you advertise the following posts which will come into effect from October 1997.
Thanks, Rik Brydson
************************************************************** School of Process, Environmental and Materials Engineering, University of Leeds, U.K.
Research Opportunities in Materials Characterization
Two positions are currently available:
o Earmarked EPSRC Ph.D. Studentship in Materials Characterization - Analytical Electron Microscopy and/or Surface Analysis applied to a wide range of materials (carbons/ ceramics/ electroceramics/ catalysts/ non-ferrous alloys and steels). Candidates should possess a 2.1 degree or higher in either Materials, Physics, Chemistry or related discipline.
o 3 Year EPSRC funded Postdoctoral Position on the modelling of electron energy loss near-edge fine structure (unoccupied electronic structure). Candidates should possess a Ph.D. in the physical sciences or an engineering discipline and have experience in two or more of the following: solid state physics/chemistry, electron microscopy and/or computing/programming.
For further details concerning both posts please contact: Dr Rik Brydson, Electron Optical Unit, Materials, Leeds LS2 9JT, U.K. (email: mtlrmdb-at-leeds.ac.uk/ tel: 0113 233 2369) ************************************************************** _____________________________ Dr. Rik Brydson, University Research Fellow, Electron Optical Unit, Department of Materials, School of Process, Environmental and Materials Engineering University of Leeds, Leeds LS2 9JT, U.K.
I am looking for an electrolyte in order to prepare some TEM thin foils of Al-Mg alloys. I have the following constraints for the eloctrolytic polishing solution : -polishing at room temperature. -solution that does not attack copper.
In addition what is the influence of the metal of the cathod (Platinum, stainless steel, copper, ...) on the result of the electrolytic polishing?
Frederic Basson McMaster University Department of Materials Science and Engineering 1280 Main Street West Hamilton, Ontario L8S 4L7, Canada Tel : (905)-525-9140 Ext 24862 Fax : (905)-528-9295 Email : bassonf-at-mcmail.CIS.McMaster.CA
We are using Tousimis otl/water filter now. The information on filter is:
Replacement Element for oil/water filter Cat. #8782A 2211 Lewis Ave. Rockville MD 20851 Tel: #301-881-2450
Wang
On Sat, 31 May 1997, Donald Lovett wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } On Fri, 30 May 1997, Steve Barlow wrote: } } } any suggestions where I can buy a water trap for my liquid carbon dioxide } } tank attached to my CPD? } } } At one time Tousimis (spelling?) sold an in-line unit. I saw the set up } at the EM facility of University of Wisconsin -at- Madison. } } } } ______________________________________________________________________ } Donald L. Lovett e-mail: lovett-at-tcnj.edu } Assoc. Professor, Dept. of Biology voice: (609) 771-2876 } The College of New Jersey fax: (609) 771-2674 } Trenton, NJ 08650-4700 } } } }
We are using Tousimis otl/water filter now. The information on filter is:
Replacement Element for oil/water filter Cat. #8782A 2211 Lewis Ave. Rockville MD 20851 Tel: #301-881-2450
Wang
On Sat, 31 May 1997, Donald Lovett wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } On Fri, 30 May 1997, Steve Barlow wrote: } } } any suggestions where I can buy a water trap for my liquid carbon dioxide } } tank attached to my CPD? } } } At one time Tousimis (spelling?) sold an in-line unit. I saw the set up } at the EM facility of University of Wisconsin -at- Madison. } } } } ______________________________________________________________________ } Donald L. Lovett e-mail: lovett-at-tcnj.edu } Assoc. Professor, Dept. of Biology voice: (609) 771-2876 } The College of New Jersey fax: (609) 771-2674 } Trenton, NJ 08650-4700 } } } }
I am experiencing an imaging problem with Photoshop 3.0.
Images are acquired by scanning in TEM negs using a Polaroid Sprintscan 45 and a Power Mac 8500. Files are saved on the Mac hard drive in TIF format for IBM's. No problem opening the IBM files from the Mac hard drive, but when the files are copied onto an IBM-formatted ZIP disk, they open as segmented images on the Mac -- like someone cut the images into strips and jumbled up the order.
I can open the images from the IBM ZIP disk using NIH Image and PageMaker on the Mac but NOT Photoshop. Now, if I drag the files from the IBM ZIP disk back onto the Mac hard drive, they open properly. I also noticed this problem on IBM formatted JAZ drive as well.
What's going on and how can this be remedied? Is this an Iomega glitch?
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 ####################################################################
I ATTEMPTED TO DO RESOLUTION MEASUREMENTS USING A LAB6 FILAMENT. I WAS ABLE TO RESOLVE A FEW MOCRONS USING A TUNGSTEN FILAMENT AT VARIOUS CURRENTS, HOWEVER WITH THE LAB6 FILAMENT I HAVE HAD PROBLEMS FOCUSING THE MICROSCOPE AND HAVE NOT BEEN ABLE TO RESOLVE THE MICRON STRUCTURES ON THE TEST SAMPLE. DOES ANYONE HAVE A REASONABLE EXPLANATION ? THE SEM IS A ISI SX40.
I am in charge of our EM facilities and have been asked to compare our charge schedule with other lab's. We have a Philips 201 and a 420,as well as an SEM. We do all types of specimen preparation (including freeze etching and cryosectioning) and dark room work. Thanks for any info which can be directed to me at the adress above or to revelj-at-cco.caltech.edu JP Revel.
} I ATTEMPTED TO DO RESOLUTION MEASUREMENTS USING A LAB6 FILAMENT. I WAS } ABLE TO RESOLVE A FEW MOCRONS USING A TUNGSTEN FILAMENT AT VARIOUS } CURRENTS, HOWEVER WITH THE LAB6 FILAMENT I HAVE HAD PROBLEMS FOCUSING } THE MICROSCOPE AND HAVE NOT BEEN ABLE TO RESOLVE THE MICRON STRUCTURES } ON THE TEST SAMPLE. DOES ANYONE HAVE A REASONABLE EXPLANATION ? THE SEM } IS A ISI SX40.
If you do not have good enough vacuum, chromatic aberration will effectively enlarge the spot size and destroy your resolution.
Also, the gun circuitry may need to be modified to permit you to effect good emission currents. Was your filament installed by service personnel and the scope properly set up to accept the filament?
#################################################################### 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 expect that the resolution change has nothing to do with that change-over, rather it points to a contamination/ stigmatism problem. Also, because of the greater brightness from the LaB6 you would probably use the condensor more focused and the smaller probe size should actually improve resolution. It is assumed that other vital working parameters (distance, kv, beam current, specimen type) are unchanged. 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 ---------- } From: Bruce Brinson {brinson-at-rice.edu}
} Subject: EM LAB6 CONVERT. } Date: Tuesday, 3 June 1997 6:08 } I ATTEMPTED TO DO RESOLUTION MEASUREMENTS USING A LAB6 FILAMENT. I WAS } ABLE TO RESOLVE A FEW MOCRONS USING A TUNGSTEN FILAMENT AT VARIOUS } CURRENTS, HOWEVER WITH THE LAB6 FILAMENT I HAVE HAD PROBLEMS FOCUSING } THE MICROSCOPE AND HAVE NOT BEEN ABLE TO RESOLVE THE MICRON STRUCTURES } ON THE TEST SAMPLE. DOES ANYONE HAVE A REASONABLE EXPLANATION ? THE SEM } IS A ISI SX40. } } THANK YOU
Email: cesa-at-compass.com.ph Name: Christian Eric S. Abaya
School: University of the Philippines
Question: I am a graduate student of the University of the Philippines, and I } am now starting doing my masteral thesis which is somehow related on } electron microscopy of microorganisms (local isolates-bacteria, fungi and } actinomycetes). I am experiencing some problem isolating and preparing a } pure strain for } TEM and SEM observation. Can you provide me some basic technique in } isolating and processing microorganism (for TEM and SEM). } Your immediate response will be very much appriciated. Thank you } very much.
From experience I could say following - as in serviced by me Philips microscopes it is possible nicely to visualize the so called beam crossover which is showing image of the filament spot after Wehnelt - I observed following: - the Lab6 is more sensitive to Wehnelt - tip distance adjustment and tip centering as W. - close-to-saturation-point looks like malthese cross with 4 bright arms not like the torus by W-filament - if the Wehnelt distance is not good you can see on image status like saturation (no more light, by more heating) but you are on one of the arms or in the middle but arms are still bright - this causes the image to be not sharp - like the shadows of soccer players on the night-illuminated stadion. - if centering is wrong - you can have by saturation even to spots (one from arm one from centertip) - by not-centered W-filament you have banana shape which anyhow can reach spot-saturation. - also if you just exchanged filament - maybe you can not reach saturation value of the heating current ? - did this microscope ever worked with Lab6 ? kind regards Krzysztof M. Herman PHILIPS E.O. Service Poland 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
} Has anyone a suggestion on how to prepare a metal sample to get a strong } electron channeling effect?
I presume you're intending to look at the specimen on an SEM? I'd think that something with nice, large crystals would make a good specimen - grind and polish a flat surface for the channeling. Perhaps a brass would work?
} I ATTEMPTED TO DO RESOLUTION MEASUREMENTS USING A LAB6 FILAMENT. I WAS } ABLE TO RESOLVE A FEW MOCRONS USING A TUNGSTEN FILAMENT AT VARIOUS } CURRENTS, HOWEVER WITH THE LAB6 FILAMENT I HAVE HAD PROBLEMS FOCUSING } THE MICROSCOPE AND HAVE NOT BEEN ABLE TO RESOLVE THE MICRON STRUCTURES } ON THE TEST SAMPLE. DOES ANYONE HAVE A REASONABLE EXPLANATION ? THE SEM } IS A ISI SX40. } } THANK YOU
If the filament isn't set up correctly in the gun, it will generate a large fuzzy spot. It may be that you just need to increase the filament current to get proper saturation - but be careful not to blow the filament. This assumes that the gun circuitry is capable of driving LaB6 and that you have a good vacuum.
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I have been asked by Dr. John E. Johnson, Jr., Editor-in-Chief of Microscopy Research and Technique (MRT), to solicit selected articles addressing recent advances in our understanding of Silver Halide Photographic Processes. The invited articles would be the basis for a topical issue of MRT entitled "Microscopic Research of Silver Halides and Related Dispersed Systems", on which I would serve as guest editor. Research in this field may give a number of excellent examples of successful solution of highly diverse and complicated tasks of imaging science of more than 150 years history and modern high technology focused to the next century. The term "microscopy techniques" may cover all known methods of light microscopy, electron microscopy (stationary and scanning methods), scanning probe techniques, image analysis and so on.
In this regard, I am writing to invite you, either alone or in conjunction with a collaborator(s) of your choice, to submit articles discussing some of the following topics concerning methodology, sample preparation and applications of microscopy in studies of silver halides and related dispersed systems (small particles and clusters of metals and metal sulfides, polymer gels and latexes, dyes, etc.):
1. Microscopic insight to mechanisms of nucleation and growth of silver halide crystals of photographic emulsions and model systems. 2. Crystalline and defect structures and phase composition of silver halides: impact of microscopy. 3. Structural and analytical characterization of silver halides and related dispersed systems by imaging and spectroscopic microscopy techniques. 4. Microscopy and ultramicroscopy studies of mechanisms of chemical and spectral sensitization and latent image formation. 5. Development and processing of silver halide photographic systems as studied by microscopy methods.
A manuscript length limitation is set at about 30 double-spaced typewritten pages, plus micrographs. An Abstract should be included. Each manuscript will be refereed, and therefore will be suitable for inclusion in Grant Proposals or Renewals. The MRT Instructions to Contributors (enclosed) should be used as a guide when preparing the manuscript. Note that the journal has a format of 8 1/4" by 11", and the figure size is 6 3/4" wide by a maximum of 9" high for a full page width figure, or 3 5/16" wide by a maximum of 9" high for a single column width figure. Therefore, your figures should be trimmed to these dimensions if you wish them to be reproduced at the same size as the originals. There is a charge to authors for color figures, but no charge for black and white figures, regardless of the number. You would need to include copyright release forms signed by the publisher for any previously published figures. Please examine MRT for examples of what we are looking for in topical papers. If you would be willing to participate in this venture, I would appreciate knowing your intentions by 1.07.97. I would need your Article Outline (with approximate number of pages and figures) by 15.09.97., and I would, at that time, make available to you a list of other contributors and article titles. The completed manuscript would be due in my office by 1.11.97. All manuscripts are peer reviewed and may require revision. The topical issue will be published within 4-6 months (this is part of our agreement) after the manuscripts are received at the publisher's office in New York (John Wiley). I hope you will agree to contribute to what we believe will be a fascinating and beneficial update of current knowledge in Silver Halide Photographic Processes.
Sincerely,
Vladimir Oleshko ********************************************************** V.P. Oleshko, Ph. D. e-mail:oleshko-at-uia.ua.ac.be Micro-and Trace Analysis Centre Tel.:+32-3-820.23.64 Chemistry Department FAX :+32-3-820.23.76 University of Antwerp (UIA) Universiteitsplein 1 Antwerpen-Wilrijk B-2610 Belgium ***********************************************************
John, Do I understand you to say that when you read from the ZIP into MAC photoshop the image comes up in jumbled strips, but when you drag the image from the ZIP to the hard drive then the image reads in correctly? If that be the case, I would wonder if there is something wrong in the ZIP drivers or hardware.
On the other hand, if you only see the problem reading into PC Photoshop from the ZIP, then I would wonder if the problem is elsewhere.
Some years ago, I did a TIF converter for our KEVEX images. I had a dickens of a time trying to please all of the TIF readers out there. There were so many options available for TIF files and the readers did not handle all of the tags correctly. I checked PC Word, PageMaker and a few other readers and got various results. Some were much easier to please than others. And when I had them 'happy', some MAC applications had problems with my format.
Since then, TIF writers and readers have undoubtedly improved so that they support more of the standard correctly, but there still could be problems depending on the version.
Off hand, I wonder if your images are being stored in strips (as is often the case) and that the strip addresses are getting jumbled. Sometimes there are advanced options for saving the images that allow you to specify the strip size, e.g., 8K or 32K; I wonder if that would have an impact on what you see. Also, are you able to save it to any other image formats or to turn off the strip option? I don't think that either GIF or Windows BMP formats use strips.
If you still have problems, you might try sending an attached image to me (not to the list) or tell me where I can access one and I can take a look at the internals and see what I can see.
At 01:51 PM 6/2/97 -0600, you wrote: } } Calling all imaging gurus, } } I am experiencing an imaging problem with Photoshop 3.0. } } Images are acquired by scanning in TEM negs using a Polaroid Sprintscan 45 } and a Power Mac 8500. Files are saved on the Mac hard drive in TIF format } for IBM's. No problem opening the IBM files from the Mac hard drive, but } when the files are copied onto an IBM-formatted ZIP disk, they open as } segmented images on the Mac -- like someone cut the images into strips and } jumbled up the order. } } I can open the images from the IBM ZIP disk using NIH Image and PageMaker } on the Mac but NOT Photoshop. Now, if I drag the files from the IBM ZIP } disk back onto the Mac hard drive, they open properly. I also noticed } this problem on IBM formatted JAZ drive as well. } } What's going on and how can this be remedied? Is this an Iomega glitch? } } Thanks, } #################################################################### } John J. Bozzola, Ph.D., Director ---------------------------------------------------- Warren E. Straszheim 270 Metals Development, Ames Lab/ISU, Ames IA, 50011 Phone: 515-294-8187 FAX: 515-294-3091
We have a refurbished jsm 35C for sale at the present time for $13,750. If interested please give us a call at 770-448-3200. We need to make room for other instruments now. Talk to Mark Rigler when you call. Thanks
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Has anyone a suggestion on how to prepare a metal sample to get a strong } electron channeling effect?
Usually an electropolished sample gives a strong electron channeling contrast I know this works for steels, iron, Al, CuAg alloy and many others.
}
Dr. Ijaz A. Rauf Department of Physics, University of Alberta, Edmonton, Alberta, Canada, T6G 2J1.Ph:(403)492-3041 Fax:(403)492-0714 E-mail:irauf-at-phys.ualberta.ca **** Love For All ** Hatred For None **** I Express My Opinions Only *** ************************************************************************ * Ahmadiyyat, in fact, is the true Islam revealed to Muhammad (PBUH) * * You can watch for yourself on satellite TV, for details about time * * and channels in your region visit URL http://alislam.org/mta/ * ************************************************************************
} Klaus } =20 } The Biomaterials Science Group } Department of Oral and Dental Science=20 } in co-operation with the Physics Department } University of Bristol } Lower Maudlin Street, Bristol, BS1 2LY, England } =20 } =20 } =20 } For the project } =20 } "Protein-Biomaterials-Interfaces Investigated with Atomic=20 } Force Microscopy" } =20 } we are looking for } =20 } a PhD Student } =20 } with a background in Physics or Materials Science or Chemistry or Biolo= gy } or Engineering } =20 } or a student of related areas. } =20 } We expect: } =B7 EU citizenship (non-EU citizens can be considered if they } provide funding which covers the difference between the=20 } oversees fee and the home fee) } =B7 Upper Second Class degree or better } =B7 Enthusiasm for the work in the area of biomaterials interfaces } =B7 Eagerness to explore new areas, initiative and result oriented work } =B7 Starting date 1st October 1997 } =B7 Open, cooperative character for the work in an international team } =20 } We offer: } =B7 Research at one of the leading British research universities in att= ractive surroundings } =B7 Top departments } =B7 PhD fees are paid } =B7 Maintenance and travel grant (co-operation with a university in Cal= ifornia possible) } =B7 Applied scientific research on a high level } =B7 Intensive supervision } =B7 Work in international teams with an excellent working atmosphere } =B7 Future oriented research area } =B7 Premium (=A3 2000) from industrial sponsor paid upon successful com= pletion of PhD research programme } =20 } Closing date for applications 15th June 1997 } =20 } Applications should be submitted to: } =20 } Dr. Klaus D. Jandt =09 } Senior Lecturer } Dental Materials Science and Biomaterials } University of Bristol =09 } Department of Oral and Dental Science =09 } Lower Maudlin Street =09 } Bristol, BS1 2LY, UK =09 } Phone: ++ 44 (0)117 9 28 44 18 } Internet: K.Jandt-at-bris.ac.uk } =20 } ----------------------------------------- } Dr. Klaus D. Jandt } Senior Lecturer } Dental Materials Science and Biomaterials } University of Bristol } Department of Oral and Dental Science } Lower Maudlin Street } Bristol, BS1 2LY, UK } Phone: ++ 44 (0117) 9 28 44 18 } Fax: ++ 44 (0117) 9 28 47 80 } Internet: K.Jandt-at-bris.ac.uk } WWW: http://mail.bris.ac.uk/~omkdj/ } =20 } =20
On 06/02/97 19:12:06 you wrote: } } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Our Safety officer is concerned about us decanting the liquid nitrogen from our dewar into our coldstage using a styrofoam cup. Does anyone have any suggestions on where we might purchase a cryo ladle of some sort. The cryo gloves that we have, are too cumbersome when using such a small styrofoam cup for the small quantity that we decant at a time.
Any suggestions would be greatly appreciated.
Susan
Susan Carbyn Atlantic Food and Horticulture Research Centre Agriculture and Agri-Food Canada Kentville, Nova Scotia B4N 1J5 Canada
A colleague would appreciate input on chemical etching Ge(111). He has used HF10%+HNO3 90% for Ge(001) and obtained nice thin regions around the hole, but for Ge(111) got only pinholes with this solution even when cutting the strength several times.
Any suggestions?
W. Sinkler
++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ Wharton Sinkler PhD Department of Materials Science and Engineering Northwestern University 2225 North Campus Drive Evanston, IL 60208-3108 tel: (847) 491-7809 fax: (847) 491-7820 email: sinkler-at-apollo.numis.nwu.edu
I've finished my senior research project and have an abstract for the MSA. Is there a specific format I should follow, or should I just attatch it as a Word document?
Thanks so much,
Erik Pauls
********************************************************* Cretaceous liability: Tyrannosaurus wrecks. Desmostylus: one weird mammal.
Erik Pauls P.O. Box 4960 Berkeley CA 94704 erik-at-uclink.berkeley.edu (510) 528-1945
I have been discussing an immunolabeling problem with Geoff McAuliffe. The original post was on the Microscopy Listserver a few days ago. The basic problem is one of double immunolabeling with mouse monoclonals.
The following message gives additional details on the labeling procedure. Since the original post, I have seen one response that suggested introducing a saturating step with fab fragments after the first labeling run.
If you have any suggestions or thoughts on other things to try, please correspond directly with Geoff at the address in the header.
Many thanks, and greetings from sunny (and HOT!) Arizona.
Bob Chiovetti (RCHIOVETTI-at-aol.com) --------------------- Forwarded message:
Dear Bob:
Thanks for responding to my querry. I should have been more specific re the objects of my study. Mouse CNS, light microscopy, paraformaldehyde fixation, 20 micron frozen sections, stained free-floating with mouse monoclonal anti Mac-1 (macrophages and microglia) with a Vector ABC Elite kit (biotinylated secondary, etc.). Mount sections on slides stain with mouse monoclonal anti-bromodeoxyuridine after trypsin unmasking (hence mounting on slides) also with a Vector ABC Elite kit. Done individualy the results are fine but when combined the staining with the second AB is greatly reduced and very spotty. The trypsin is essential after formalin fixation, and Mac1 does not survive any fixation that allows omission of trypsin (alcohol). I don't think I can do the Bromodeoxyridine first since the trypsin will digest the cell surface marker Mac-1 detects. Hot buffer antigen retrieval does not give good results with bromodeoxyuridine. If I use a rabbit polyclonal for GFAP at the first antibody I get beautiful double staining but of two seperate populations of cells so I am not sure if my problem is trying to use two mouse monclonals to stain two things in one cell or two things on one section. I am getting the idea that the secondaries are the problem? so perhaps one or both primaries linked to gold? I do need to see two colors, though. Thanks in advance!
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 ***************************************************************
We are setting up a fluorescence microscopy lab. Would like info on insect tissue preps. lisa & vickie Vickie A. Kimler, Ph.D. Biology and Allied Health Department Mercyhurst College Erie, PA 16546 1-814-824-2169
} Our Safety officer is concerned about us decanting the liquid nitrogen } from our dewar into our coldstage using a styrofoam cup. Does anyone } have any suggestions on where we might purchase a cryo ladle of some } sort. The cryo gloves that we have, are too cumbersome when using } such a small styrofoam cup for the small quantity that we decant at a } time. } } Any suggestions would be greatly appreciated. } } Susan
This subject has been discussed a number of times before - it seems to be a common experience that safety officers don't have much/any experience of LN2 and tend to come up with all sorts of concerns which are generally wrong.
For example, gloves - they are cumbersome and make handling difficult, so you are more likely to spill the LN2. If it goes inside the gloves, you're in real trouble. Whereas some LN2 spilling on to bare skin will cause no damage at all.
First I would ask your safety officer to clarify precisely the concern about the way you are currently handling the LN2. As I say, a small spill on to bare skin will cause no damage - if you slowly pour several litres over your hands, then you might get a burn.
I would also bring up the question of shoes. If you are wearing shoes and spill LN2 into the shoe, you'll get a nasty burn. If you are not wearing shoes or socks, there will be no problem (unless you insist on standing in a puddle of LN2). So, logically, when handling LN2, all shoes (and socks) should be removed:) From personal experience, I would even suggest that handling LN2 is best done completely naked - if it spills on to your clothing, it'll get held against the skin and cause a burn.
In your case, if the safety officer insists, I'd get a strip of aluminium and bend it around to make a handle with a loop at the end where the styrofoam cup can sit.
I'm not against safety officers - it is an important job which needs doing. But enforcement of regulations, possibly written to cover rather different circumstances, by people who don't fully understand the real risks (for example, with LN2 large amounts of N2 gas are generated - has proper ventilation been considered) is not the right approach - it leads to people ignoring safety officers and saftey rules.
I'm not familiar with the kits you're using, however it appears that you're staining with a mouse primary followed by an anti mouse reagent followed by another mouse primary, is that right? If it is then there's a likelihood that the free arm/s of the second layer attached to the cells via the first primary are capturing the second primary when you put it in. You can prevent this cross-linking by using a FAB second layer, or more cheaply, if the second primary is directly conjugated (or biotinylated), you can block the spare arms of the antimouse by adding normal mouse serum after the first second layer and before the second primary.
Ray
At 8:33 pm -0400 3/6/97, RCHIOVETTI-at-aol.com wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Ray Hicks ________________________________________________________________________ |University of Cambridge |Tel 01223 330149 | |Department of Medicine |Fax 01223 336846 | |Level 5, Addenbrookes Hospital |e-mail {rh208-at-cus.cam.ac.uk} | |Hills Road Cambridge |Web http://facsmac.med.cam.ac.uk | |CB2 |ftp server ftp://131.111.80.78 | |UK | | |_________________________________|_____________________________________|
On Wed, 4 Jun 1997 08:09:03 +0100 Larry Stoter {LPS-at-teknesis.demon.co.uk} wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } Our Safety officer is concerned about us decanting the liquid nitrogen } } from our dewar into our coldstage using a styrofoam cup. Does anyone } } have any suggestions on where we might purchase a cryo ladle of some } } sort. The cryo gloves that we have, are too cumbersome when using } } such a small styrofoam cup for the small quantity that we decant at a } } time.
} This subject has been discussed a number of times before - it seems to be a } common experience that safety officers don't have much/any experience of } LN2 and tend to come up with all sorts of concerns which are generally } wrong. } } For example, gloves - they are cumbersome and make handling difficult, so } you are more likely to spill the LN2. If it goes inside the gloves, you're } in real trouble. Whereas some LN2 spilling on to bare skin will cause no } damage at all. } } First I would ask your safety officer to clarify precisely the concern } about the way you are currently handling the LN2. As I say, a small spill } on to bare skin will cause no damage - if you slowly pour several litres } over your hands, then you might get a burn. } } I would also bring up the question of shoes. If you are wearing shoes and } spill LN2 into the shoe, you'll get a nasty burn. If you are not wearing } shoes or socks, there will be no problem (unless you insist on standing in } a puddle of LN2). So, logically, when handling LN2, all shoes (and socks) } should be removed:) From personal experience, I would even suggest that } handling LN2 is best done completely naked - if it spills on to your } clothing, it'll get held against the skin and cause a burn. } } In your case, if the safety officer insists, I'd get a strip of aluminium } and bend it around to make a handle with a loop at the end where the } styrofoam cup can sit. } } I'm not against safety officers - it is an important job which needs doing. } But enforcement of regulations, possibly written to cover rather different } circumstances, by people who don't fully understand the real risks (for } example, with LN2 large amounts of N2 gas are generated - has proper } ventilation been considered) is not the right approach - it leads to people } ignoring safety officers and saftey rules. } } Regards, } Larry Stoter } } I have the misfortune to be the safety officer in my deparment and I would like to endorse just about everything that Larry Stoter has to say about the handling of small volumes of liquid nitrogen. Eye protection in the form of goggles or a face shield is highly desirable, so total nudity is not a good idea.
Regards, Eric Lachowski
---------------------- Dr Eric E. Lachowski University of Aberdeen Department of Chemistry Meston Walk Old Aberdeen AB24 3UE +44 1224 272934 e.lachowski-at-abdn.ac.uk
On the subject of LN2, does anyone know the correct first aid treatment for a burn from this substance?
Normally, one would use cold water to cool a (heat) burn and prevent further tissue damage. But that seems inappropriate somehow...Hot water?
Perhaps in view of the known and imagined dangers and the lack of medical information re-treatment we should all wear a full immersion suit and breathing aparatus to enter any building where LN2 is stored. -- Anthony James Bentley Surface Data Scientific Instrumentation and Software Web site http:\\www.surface.demon.co.uk
I am pleased to see the thread on liquid nitrogen safety. Our lab does a lot of cryoTEM so we use lots of liquid nitrogen. Like Larry Stoter, we prefer no gloves and open safety glasses so any liquid nitrogen that might bounce onto the skin can escape easily. I was able to convince our safety officer that this made sense. We have one exception: we handle glass wide-mouth dewars (often with steel jackets.) We had one implode and send a shower of glass through the lab with such force that fragments were embedded in a fabric chair cushion. When we fill these wide-mouth glass Dewars, we wear a full face shield. -- 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
It probably is a function of the crystal orientation because 111 typically etches into tetrahedral holes while 100 etches with flat bottomed holes. I think if you consult some of th eearly literature you will find the best etch.
A colleague would appreciate input on chemical etching Ge(111). He has used HF10%+HNO3 90% for Ge(001) and obtained nice thin regions around the hole, but for Ge(111) got only pinholes with this solution even when cutting the strength several times.
Any suggestions?
W. Sinkler
++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ Wharton Sinkler PhD Department of Materials Science and Engineering Northwestern University 2225 North Campus Drive Evanston, IL 60208-3108 tel: (847) 491-7809 fax: (847) 491-7820 email: sinkler-at-apollo.numis.nwu.edu
Liquid nitrogen burns would be treated as frostbite. There are several degrees of frostbite depending on how deep the freezing damage is. Most exposure is acute, a splash on the hand or arm. The liquid nitrogen evaporates quickly enough that it cause at the most a slight reddening of the skin similar to a sunburn. This just requires observation of the sight to see if any further blistering occurs. A more severe case would involve submersion into the liquid nitrogen. This would cause more deeper damage. You must keep the frozen appendage still, in the case of hands and fingers (toes or feet) they can be rewarmed by placing in a pan of warm (~102 F) water, or under your arm. Wrap the damaged part in a sterile dry gauze, there will be blistering-do not break open the blisters, and seek immediate medical attention. If there is any doubt about the level of injury you should seek medical attention. This information is from the National Ski Patrol-Outdoor Emergency Care, and is the method of treatment for frostbite we teach to patrolers. Linda Barthel Research Associate II Department of Anatomy and Cell Biology University of Michigan lab (313) 764-7476 fax (313) 763-1166 barthel-at-umich.edu
and National Ski Patrol Outdoor Emergency Care Instructor, Mt. Brighton Michigan.
Susan Get a very small dewar, we use a 1 liter dewar for small transfers, keeps the safety people happy. Do not use a regular vacuum bottle (Thermos) as it can break and be more dangerous then the styrofoam cup. Also the plastic cryo dewars can break we broke two so far this year. I would recommend metal.
Ask the Safety person where to get a small dewar he should have the necessary catologs with part numbers. Then the safety people know you are interested in listening to them. Ours is made by Aladdin Ind.
our 4 liter is made by Union Carbide. Your LN2 supplier may also have these.
Keith Collins Albany Research Center Department of Energy Albany, Oregon collins-at-alrc.doe.gov
One person's two-pennyworth re. liquid nitrogen, cryogens etc:
I would personally be far more concerned about boiling water than boiling LN2, bearing in mind things already said on the list about the hazards of LN2 splashes on bare skin. Those who know me or who have attended the Seefeld-in-Tirol cryo-workshops will have seen my 'drinking' of the substance.
It is true that safety officers normally know very little about the realities of the LN2 and its handling, but they have a serious job to do (I know, I am also one!).
There is a huge difference between primary coolants i.e. those liquids which boil at very low temperatures (LN2 and LHe) and secondary coolants which have to be cooled by LN2 such as liquefied ethane and liquefied propane.
The secondary coolants will stick to the skin/eyes and boil off at c. -80 (ethane) and c. -40 (propane), in other words your body has to heat them up e.g. 150 degrees C in the case of propane to turn it into a gas. As body temperature is about 40 degrees C, this becomes a problem. With these, this boy does take the precautions (full face mask while liquefying and disposing), but with 'thinnish' leather gardening gloves so as to retain handling sensitivity (otherwise you are into a dangerous practice!). The thick cryo-gloves in this lab are retained solely for handling the delivery hose on the big LN2 storage tank.
A far greater risk from using LN2 is that of ASPHYXIATION resulting from poor ventilation, bearing in mind that each litre of LN2 forms approx 690 litres of gaseous N2 depending on ambient temperatures. This does tend to dilute the 18% (?) oxygen concentration in normal air somewhat. In this respect, I know of one very close encounter of the terminal kind.
In these situations I often cite "Safety consideration regarding the use of propane and other liquefied gases as coolants for rapid freezing purposes" by KP Ryan & MI Liddicoat (1987) J. Microsc. 147, 337-340.
Claimer - I wrote it.
With best wishes - Keith Ryan
++++++++++++++++++++++++++++++++++++++++++++++++++ Keith Ryan B.Sc., Ph.D., C.Biol., M.I.Biol. Plymouth Marine Laboratory, Citadel Hill, Plymouth, Devon PL1 2PB, England
My approach to this has been to be sure there are lots of open safety glasses around and to use one of those plastic beakers. You can get them with handles or, what we've done, is to save those handles from broken coffee carafes. They will last years or until some moron slams them down on a hard surface whilst cold. I've seen people trying to carefully decant lN2 from a wide mouth Dewar and it looks less safe. I've also seen people demonstrate the relative safety of lN2 by quickly dunking and removing their hand from a Dewar. It might convince your Safety Officer of something...
cheers, John Heckman Michigan State University Center for Electron Optics
Our Safety officer is concerned about us decanting the liquid nitrogen } from our dewar into our coldstage using a styrofoam cup. Does anyone } have any suggestions on where we might purchase a cryo ladle of some } sort. The cryo gloves that we have, are too cumbersome when using } such a small styrofoam cup for the small quantity that we decant at a } time. } } Any suggestions would be greatly appreciated. } } 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
Does anyone have on-line sign-up for equipment time? I remember this was discussed here ages ago, but I haven't heard about it recently. We are thinking about going to such a system, and would like to hear pros/cons/horror stories/suggestions.
........."Those who know me or who have attended the Seefeld-in-Tirol cryo-workshops will have seen my 'drinking' of the substance. "......
Yes, indeed I do recall the 'smoking ears and nose of Keith' during the workshops.
I have been using this substance since 1964 in large quantities. The best approach is common sense, that is what I have always demonstrated and taught to the many new comers in the exiting field of cryo applications.
Beware of plastic funnels. For several years I used a high density polyethelene funnel during some LN2 transfers. One day while being used, the HDPE exploded without warning. Shards of funnel flew all about the lab. Never did an examination to determine the cause, but suspect that over the years, micro-cracks had developed and at some point the thermally induced stress/strain was more than the (remaining) cold embrittled HDPE could stand.
I would suggest using funnels of teflon or stainless steel. ... Or none at all as is now my practice- not difficult to clean-up the spills {g} .
Responding to the message of {199706041753.NAA27849-at-phage.cshl.org} from howard-at-cshl.org (Tamara Howard): } } Does anyone have on-line sign-up for equipment time? I remember this was } discussed here ages ago, but I haven't heard about it recently. We are } thinking about going to such a system, and would like to hear } pros/cons/horror stories/suggestions. } yes, we have a web-based sign up based on our FileMaker Pro database. You can see what it looks like from our web site at URL http://resolution.umn.edu.
(It is linked as "Instrument sign-up") You will not be able to sign up (or even see the actual sign-up form) unless you have a username and ciode number here. This is a useful security feature to prevent millions on random surfers from booking all the time on our instruments.
I am currently trying to migrate to a less me-specific platform than the kludgy applescript that now does all the hard work.
This system has grown over the years and was not instantly developed as the giant behemoth it now is.
You will need to think about what size blocks of time you want, whether people can cancel their time and when, or whether they can delete other peoples time. I know it shouldn't be necessary, but have had it proved otherwise :(
Stuart
__________________ Stuart McKernan stuartm-at-tc.umn.edu Microscopy Specialist CIE Characterization Facility, University of Minnesota Phone: (612) 626-7594 100 Union Street S. E., Minneapolis, MN 55455 Lab: (612) 624-6590
To all list members on behalf of Richard Easingwood:
} To: richard.lander } From: Richard Easingwood {richard.easingwood-at-stonebow.otago.ac.nz} } Subject: Glutaraldehyde: safe limits
I originally posted this message to a safety listserver where there is a discussion running regarding safe limits for glutaraldehyde and glutaraldehyde monitoring (so it starts in mid 'discussion'). Somebody suggested I post this on this list as well, I think it is probably appropriate to anyone using GA, and I would certainly be interested to hear any feedback. From a personal point of view I'd be particularly interested to hear from anyone who thinks they've become sensitised to GA and what steps they take to minimise exposure.
} } I too am very skeptical about the maximum 'safe' exposure limit of 0.2ppm, } I am also doubtful of the use of the Glutaraldemeter (mentioned in one of } the previous replies in this discussion) for measuring safe limits as a } result. I should mention at this point that I have developed a sensitivity } to glutaraldehyde (GA), probably as a result of a small number of slight } exposures to this chemical over the last 6 years in my field of electron } microscopy. I know that if I exposed myself to air contaminated to 0.2ppm } (as measured by the Glutaraldemeter) it would be dramatically } debilitating for me - the 'steel band' goes around my chest, my energy } levels drop to nothing and I get depressed (common symptoms for this } ailment) The depression is so acute it is obviously something chemically } induced, as easy to pin point the cause as are jitters after too much } coffee. Takes about 12 hours before I feel normal again. } Now, I realise that most people suffer no such symptoms upon exposure but } neither did I for the first 5 years and I was very careful. Glutaraldehyde } was known to be hazardous when I first started using it in 1990 and this } was emphasized to me. In all the time I've used it I've actually only } smelt it a handful of times. Now I can only use our preparation lab when a } suitable mask, if I get a whiff of GA its too late for me. } } Yesterday we measured (using a Glutaraldemeter) the GA concentrations in } the air during various laboratory procedures (some very sloppy on purpose) } and the levels at no stage went higher than 0.15ppm. The smell of GA was } very strong during some of these as judged by two other volunteers (not } me, I wore my mask). Our OSH guidelines state that the odour threshold for } GA is about 0.04ppm which does not tally with our measurements made with } the meter, the smell was gauged as 'very strong' when the meter read below } this level. The meter was recalibrated before the tests and checked again } immediately afterwards. I can send a report to anyone interested. } I should say I have no desire to run down the Glutaraldemeter. We were } using it close to its limit of sensitivity afterall. I think badges fall } into the same category - they are probably fine if you want to ensure you } don't go over the official limit, the problem is that I think the limit is } too high. } } In conclusion I would say that the official safe maximum peak exposure } level of 0.2ppm is too high if you want to avoid the risk of becoming } sensitised to GA (and all the inconvenience that brings). I would say that } if you can smell it, the level of GA in the air is too high. I know that } this probably sounds hopelessly impractical for those working in Hospitals } where GA is in widespread use but I believe that we are only starting to } appreciate how dangerous GA is. } } Regards, } Richard } }
Richard Easingwood South Campus Electron Microscope Unit School of Medical Sciences University of Otago PO Box 913 Dunedin NEW ZEALAND
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } I would also bring up the question of shoes. If you are wearing shoes and } spill LN2 into the shoe, you'll get a nasty burn. If you are not wearing } shoes or socks, there will be no problem (unless you insist on standing in } a puddle of LN2). So, logically, when handling LN2, all shoes (and socks) } should be removed:) From personal experience, I would even suggest that } handling LN2 is best done completely naked - if it spills on to your } clothing, it'll get held against the skin and cause a burn. } I agree with your comment about shoes, after cycling to work on a very wet night and immeadiately going to fetch an evening's supply of LN2, I found I had to fill the tipping 25l dewar from the pressurised 100l one first. This involves clouds of vapour pouring onto the floor for several minutes and I found my feet getting cold as my damp shoes started to freeze. Bare feet were safer, but maybe if I polished the shoes more often they wouldn't have got so saturated.
Alasdair Preston Dept of Mat Sci Met Cambridge Univ Pembroke Street Cambridge UK yx10000-at-cus.cam.ac.uk
} } Our Safety officer is concerned about us decanting the liquid nitrogen } } from our dewar into our coldstage using a styrofoam cup. Does anyone } } have any suggestions on where we might purchase a cryo ladle of some } } sort. The cryo gloves that we have, are too cumbersome when using } } such a small styrofoam cup for the small quantity that we decant at a } } time. } } } } Any suggestions would be greatly appreciated. } } } } Susan _______________________________________________________ Prabath Perera, Ph.D. SWT Department of Physics Research Associate San Marcos, TX 78666 Email: pp14-at-swt.edu Phone: 512/245-2131 http://www.swt.edu/~pp14/ Fax: 512/245-8233
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List members, This message on behalf of Allan Mitchell,
Sometime ago I asked several questions on the listserver about additives to use in the closed circuit cooling systems of electron microscopes. For various reasons I wanted to revisit the additive requirements of the closed circuit cooling system on our TEM's. Below is a summary of a larger report that outlines the information I was able to obtain and the conclusions I drew.
If any one would like a copy of the full document then contact me at the email address below.
Introduction For numerous reasons a closed-circuit water cooling system is the preferred option for providing cooling water to the electron microscope. Cooling water is required by the electron microscope to cool the diffusion pumps and to keep the electronic's and column temperature stable.
A closed-circuit water cooling system is essential if the local water supply has a high chloride concentration, has floating particles, is acidic, has a water temperature that fluctuates and is uncontrollable (this potentially leads to specimen drift problems in the TEM) and / or has a water temperature that is very cold (this potentially leads to condensation problems or diffusion pumps not functioning properly in the TEM).
Closed-circuit cooling systems have many advantages which includes providing significantly better control of the cooling water temperature through 'heat-producing' equipment and are less susceptible to biological fouling (from slime and algae) and to the build up of scale deposits. Because of the small water 'top-up' requirements control of potential problems is greatly simplified.
In many areas closed systems are required by local by-laws because open systems (ie, connected to town supply) are extremely wasteful of water.
For further information about closed-circuit water cooling systems on the EM can be obtained from the following two books;
Vacuum methods in electron microscopy, by Wilbur C. Bigelow (1994). Pg 214 to 217 Volume 15 in the series Practical Methods in Electron Microscopy, editor Audrey Glauert
Design of the Electron Microscope Laboratory, by R.H. Alderson (1975). Pg 46 to 49 Volume 4 in the series Practical Methods in Electron Microscopy, editor Audrey Glauert
My problem With the installation of our new TEM (Philips CM100) I decided it was time to review the additives we had been be using in our closed-circuit water cooling systems. We have two systems, one for cooling our Akashi 002A TEM and the other system for cooling a Philips 410LS TEM and a Philips CM100 TEM. Both systems ran different additives; this was a historical situation.
My first aim was to find out a more about the need for additives in the TEM cooling system and my second aim was to settle on one additive that I could use in both of our TEM cooling systems.
There are a number of factors to be considered when choosing such an additive. Firstly, the anti-corrosion function; over the years I had heard a few horror stories about the inside of TEM lens coils "rusting' away due to the fact that no precautions had been been taken against corrosion.
Secondly, the anti-microbial requirement; stopping bugs growing in the system and clogging up filters and small water lines.
Thirdly, the additive should not be depleted to quickly nor require the regular replacement of all the water in the cooling system, the additive activity must be able to be replenished (each of our two reservoir tanks hold 160 litres of water and I didn't want to be replacing these volumes very often).
To start off I decided to seek advice from the Microscopy Listserver, it is here where I found the widest range of opinions.
Next I consulted some 'water experts'. Interestingly it turned out that those who could advice me about keeping 'bugs' out of the system could not advise me much about corrosion and vica versa.
Our Solution Through BDH I had discussions with Coalite Chemicals, the supplier of a wide range of biocides.
They recommended to me a product called Phylatol. Phylatol is an aldehyde biocide, has a pH of 7.0 and contains no metal ions. It is a broad spectrum biocide with the same activity as Panacide M but with a neutral pH. They also indicated that the pH of the water containing Phylatol could be adjusted up slightly if desired. I had indicated that I would like to use a pH of 7.5 to 8 and would like to adjust the pH with Sodium Bicarbonate.
We are going to use Phylatol at a concentration of 0.2% and make it up in filtered tap water.
We will buffer the Phylatol to pH 8.00 with Sodium Bicarbonate. This requires only a very small amount of Sodium Bicarbonate, approximately 0.3 gram in 2 litres of water.
Some other interesting points. Some other interesting points that came up in the discussions follow. The dimensions of the water channels within the electron microscope are often quite small. It is very important to exclude fibrous or particulate matter form these pipes both during assembly of the water lines and during cleaning of the reservoir tanks.
We have had personal experience of the frustration of insufficient water flow only to eventually find a lump of plumbers hemp (or something similar) lodged in a lens coil cooling line restricting the flow.
Ideally the materials of construction of a closed circuit cooling system should have smooth surfaces to resist being colonised by microorganisms. Bacteria tend to grow on a solid surface but they can not lodge onto a surface while a brisk flow of water is maintained. It is advisable to avoid areas of 'stagnant' water where possible in the reservoir. Bringing the return into the tank in such a way that promotes a swirling motion is one way of doing this.
Storage tanks should be smooth walled inside with a conical or dished bottom so that water can be drained completely at the time of water replacement .
Pumps with a small number of large vanes should be avoided and instead use a pump with a large number of small vanes, an Archimedes screw type pump being the best. This is to reduce pulsations in the water line which could potentially lead to specimen movement in the TEM.
The pump should be able to deliver a greater pressure than required as pressure drops off quickly in small lines. It is a good idea to use large lines right up to the EM.
To help eliminate pulsations it is suggested that a coil of copper pipe (with several coils) be located beside the pump and use a long run of plastic tubing rather than metal pipe to the microscope. If the problem is very serious then a rubber diaphragm system with air above the diaphragm to compress will help. If using copper tubing at all it must not connect to the microscope to prevent any earth loops.
Right angle bends in the plumbing should be avoided as these can also set up turbulence in the water which can lead to specimen movement problems.
One last but very important aspect to consider is the maintaining the correct pressure to the various pathways inside the microscope. This aspect will be the 'subject' of my next investigation.
Allan Mitchell South Campus Electron Microscope Unit School of Medical Sciences Dunedin New Zealand
We are trying to scan thinsections of rock, and are interested in obtaining a suitable scanner. The resolution should be on the order of typical 35mm film scanners ... its just that the sample isn't a typical 35mm slide. We've tried a flatbed with a transparency option, but it doesn't give the detail we want. Has anyone tried this??
Please contact me directly ...
cheerios, shAf -- {\/} /\ {\/} /\ {\/} /\ {\/} cogito, ergo zZOooOM {\/} /\ {\/} /\ {\/} /\ {\/} 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 trying to compare the distribution of particles in histological tissues and I would like to determine the distance(s) of particles relative to their nearest neighbor. Since the particles are not of uniform size I thought that if I determined the centroids it will give me a reproducible parameter. I can determine distances by triangulation but there must be a software out there that can do this faster. Does anyone know of such a software? I would appreciate comments.
Beside a full face mask when liquefying ethane, propane etc., I do insist on goggles when people are decanting from a e.g. 25 litre dewar on trunnions into the 1-litre glass dewards in steel shells (I know, we should replace these with plastic). The reason for this is a story to do with their seals: the seals at the top ens tend to be incomplete i.e. there is a gap. The anecdote, from a witness, is that in one lab some LN2 went down the gap aand blew out the glass dewar, in pieces, with some violence. Also, glass can simply fail eventually (and our glass dewars are now 16 years old and used constantly for topping-up the EDX detectors).
Another horror story: in this lab, unbeknown to me (as manager of EM plus LN2, and also Safety), a student was told bld by someone who shopuld have known better to take some LN2 in a standard picnic Thermos with a screw top. He screwed on the top and transferred to a nearby lab. It made a very good BOMB! We were picking glass out of fish tanks for days afterwards, plus all the work of transferring fish etc. DO NOT SCREW ANY LIDS ONTO LIQUID NITROGEN CONTAINERS, FOLKS! It boils back to a gas as temperature rises and if it is in defined volume then pressure rises until something gives.
Dear All I agree with what was said in this posting, except that in my case I would have to substitute formaldehyde.
The sensitisation occurred one evening in 1972. I spent several hours over a dissecting microscope working on a fish head/brain preparation after perfusion with 4% formalin + 5% glut. in a small room with no ventilation.
About 3 am I woke up with really burning eyes. I was taken to the (fortunately local) eye hospital immediatelt only to be flushed out (not nice) and given eye drops against infection. My eyes were covered for about 4 days. I couldn't go back to contact lenses for years afterwards.
I am now overpowered by low concentrations of formaldehyde. Even 0.04 ppm(measured by a meter) seems far too much too me and my eyes inflame, my throat becomes sore and sometimes my sinuses will become irritated if I stay there too long. In our lab, it is only used in a fume cabinet/cupboard or under a strong extractor fan in one place.
I would like to know why you moved to LaB6 as in my experience the SX40 will perform pretty well with W if handled correctly. What kV are you using would be of assistance in solving your problem, that is if it is no= t the LaB6 set up? For performance - 1. Working distance less than 10mm, 5 to 8 if working at {15kV 2. Spot size must be beyond half way, probably ~60% if gun is correctly set, depends on specimen emission characteristics 3. If gun is set up correctly for W emission current should be standing current plus 100uA 4. With LaB6 if filament is correctly set you should be able to obta= in an image even at the smallest spot size, if not double check your "saturation", first peak is bright but the dip following is the highest resolution position. 5. If 4 is not possible move the filament forward, current ~ standin= g current plus 20 to 50uA 6. Remember the highest resolution will only be obtained when the hi= gh voltage tank "heat gained =3D heat lost", about 1 to 11/2 hours with the= kV on! 7. Dirty column should not be a problem if you run at 30kV but lowering the kV will make the beam more susceptible to column contamination.
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Haevy metals will give more backscattered electrons than light ones. A low dislocation content and a clean surface helps also.
If you intend to do just a demo of an electron backscattering pattern EBSP or a SACP pattern , I would suggest to take the filament (W) of an old incandescence bulb which died under normal aging. Just break the glass with a glass saw (or a hammer, after embedding it in a cloth to take care of your fingers), pick the filament. You will see nice faceted W crystals due to recristallisation. EBSP/SACP patterns present a very high contrast. Better use a spot/low voltage bulb, it will have a larger filament diameter.
Regards
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 checked my "bible" and found about 80 pages of Ge etchants, not including Ge alloys. Etchants range from air to mercury nitrate.
CRC Handbook of Metal Etchants Walker, Perrin & Tarn, William H., Eds. 1991 CRC Press, Boca Raton, FL ISBN 0-8493-3623-6
1400 pages. Includes references.
------------------------------------------------ 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 --
hi Folks, we would like to use xcosm to deconvolve some z series data sets, however we need to strip the data and present it as a raw series to the package, we are able to generate tifs, picts, or other more proprietary formats (BDS DAT files, or Imagespace series) from the micrscopes but need to prepare the data for xcosm, I would like to hear how others have gone about solving this problem Tx
Simon C. Watkins Ph.D. Associate Professor and Director CBI University of Pittsburgh Pittsburgh PA 15261 tel:412-648-3051 fax:412-648-8330 URL http://sbic6.sbic.pitt.edu
--IMA.Boundary.943125568 Content-Type: text/plain; charset=US-ASCII Content-Transfer-Encoding: 7bit Content-Description: cc:Mail note part
Michael,
How about using the Polaroid 35 mm slide scanner with the histoslide adapter? We use it quite successfully to scan large histoloty sections at high resolution. Down side is time to scan and large file size. Check with Polariod.
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We are trying to scan thinsections of rock, and are interested in obtaining a suitable scanner. The resolution should be on the order of typical 35mm film scanners ... its just that the sample isn't a typical 35mm slide. We've tried a flatbed with a transparency option, but it doesn't give the detail we want. Has anyone tried this??
Please contact me directly ...
cheerios, shAf -- {\/} /\ {\/} /\ {\/} /\ {\/} cogito, ergo zZOooOM {\/} /\ {\/} /\ {\/} /\ {\/} 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/
(IMA Internet Exchange 2.1 Enterprise) id 0010A583; Wed, 4 Jun 97 19:13:02 -0500 Received: from Sparc5.Microscopy.Com (sparc5.microscopy.com [206.69.208.10]) by ns1.baxter.com (8.8.0/8.8.0) with SMTP id TAA19835; Wed, 4 Jun 1997 19:21:30 -0500 (CDT) Received: (from daemon-at-localhost) by Sparc5.Microscopy.Com (8.6.11/8.6.11) id QAA04873 for dist-Microscopy; Wed, 4 Jun 1997 16:57:05 -0500 Received: from aaem.amc.anl.gov (aaem.amc.anl.gov [146.139.72.3]) by Sparc5.Microscopy.Com (8.6.11/8.6.11) with SMTP id QAA04870 for {Microscopy-at-sparc5.microscopy.com} ; Wed, 4 Jun 1997 16:57:04 -0500 Message-ID: {3395E48A.C31C36B2-at-darkwing.uoregon.edu}
On Wed, 4 Jun 1997, Corazon D. Bucana wrote:
} I am trying to compare the distribution of particles in histological tissues } and I would like to determine the distance(s) of particles relative to their } nearest neighbor. Since the particles are not of uniform size I thought } that if I determined the centroids it will give me a reproducible parameter.
It will but the data will be quite different from what you might get from measuring the shortest distance between the outer perimeters of the objects. That, of course, is a much more tedious process. (We have faced some of the these issues before since we are interested in the size and spatial distribution of myofibrils in striated muscle.) One way to assess the distances between object, as opposed to that between centroids, is to use stereological measurements of the spaces between the objects at varying angles of testing.
PRINCIPAL DUTIES. Provide technical assistance to projects which focus on receptor mediated gene transfer into human cultured cells. Maintain human cell culture stocks under the guidance of Associate Scientist. Record microscopic images of the cultured cells. This job shall also include preparation of the media, freezing and thawing of stocks, sterilization of glassware, keeping records, maintaining supplies, general lab clean-up. The projects are described on WWW: {smaller}
EXPERIENCE REQUIRED. Hands-on experience with sterile work with cell cultures under the laminar flow is required.
PROFESSIONAL OPPORTUNITIES. There is an opportunity to become involved in preparation of cells for modern imaging technologies e.g. two-photon excitation, confocal, low-dose fluorescence with deconvolution of images, rapid cryoimmobilization, in situ PCR, immunolabeling, eftem, but maintaining cell cultures will be the primary responsibility. This is a research job. Flexible work hours possible. Equal opportunity employer - minorities are strongly encouraged to apply. {fontfamily} {param} Times {/param}
{/fontfamily} APPLICATIONS. Job will become available on the15th of June1997. Applications will be accepted until position is filled. Candidates are encouraged to submit applications, CVs, names of referees, and/or letters of recommendations to: Dr. Marek Malecki, P.I. {fontfamily} {param} Times {/param}
{/fontfamily} Address: Integrated Microscopy Resource and Molecular Biology Laboratory, rm159, 1675 Observatory Drive, Madison, WI53706. {fontfamily} {param} Times {/param}
{/fontfamily} Email: malecki-at-macc.wisc.edu {fontfamily} {param} Times {/param}
} On Wed, 4 Jun 1997, Corazon D. Bucana wrote: } } } I am trying to compare the distribution of particles in histological tissues } } and I would like to determine the distance(s) of particles relative to their } } nearest neighbor. Since the particles are not of uniform size I thought } } that if I determined the centroids it will give me a reproducible parameter.
If the distance you really want is the separation between centroids of the sections in the plane, you can get if from their centroids. If what you really want is the 3D distance between the surfaces of the particles, you can get it by drawing random lines (which are random in 3D if your section was random) on the image and measuring the length of the intercept lines across the inter-particle background. This is a tad complicated sounding, but actually rather simple. Bear with me Each intercept length we will call L. You want to determine the average value of the reciprocal, 1/L. The average value of 1/L is two-thirds of the value of 1/T where T is the actual mean interparticle spacing (surface to surface). The proof is geometrical and somewhat arcane, but well known [;-)] in the stereological literature.
One of our customers is inquiring about the possibilities for time lapse video recording and motion analysis. If you have a system that you are happy with, I would appreciate hearing about it.
Vendors are also encouraged to respond, but *please,* no sales pitches on the list. Post to the list only if the material is of an educational nature or involves new technology. Otherwise, please respond directly to me off-list.
The questions we need answered are as follows. This would be for a system which is used on an inverted microscope for cell culture:
1. Option #1 would be to equip an existing inverted microscope for time-lapse video recording. This scope already has a CCD camera on it. What are the possibilities for time-lapse VCRs?
2. Option #2 (I'm assuming) would be to interface the camera to a computer with a framegrabber board that runs motion analysis software. What is the state of the art here? Also, what are the requirements for computing horsepower?
3. Option #3: Could we combine #1 and #2? In other words, could we acquire images with a time-lapse VCR, then later send the images to the computer for motion analysis?
Your thoughts and opinions would be greatly appreciated. Thanks!
Bob Chiovetti E. Licht Company (RCHIOVETTI-at-aol.com)
I am doing a friend a favor (no good deed ever goes unpunished). He has a backscatter detector on a Hitachi SEM but has lost most of the documentation. The unit is labeled "GW Electronics Type 113." His question is "What is the resolution of the detector (smallest change in Z detectable) and is it a function of accelerating voltage?"
I'm getting some human hepatocytes for possible DAB staining within the next couple weeks. I have no experience with human hepatocytes or DAB. Can anyone recommend an appropriate fixative and buffer? Also, how long can I store these and they'll be o.k. for DAB staining? Should I store them in buffer after a period of fixation? Any help / advice would be appreciated.
Marcia Pitzenberger Merck & Co., Inc. P.O.Box 4, WP45-251 West Point, Pa 19286-0004 (215)652-9767 marcia_pitzenberger-at-merck.com
Frank wrote, } } I am doing a friend a favor (no good deed ever goes } unpunished). He has a backscatter detector on a Hitachi } SEM but has lost most of the documentation. The unit is } labeled "GW Electronics Type 113." His question is "What } is the resolution of the detector (smallest change in Z } detectable) and is it a function of accelerating voltage?" } Dear Frank, Several conditions affect the smallest Z detectable. They are KV, atomic number, tilt of the stage, beam current and scan rate. If you would like, I can fax to you part of the manual for the type 30a/113a backscattered electron detector. It is about 18 pages long. Call or e-mail me.
Gregory Rudomen University Microscopy Imaging Center S.U.N.Y. Stony Brook Greg-at-UMIC.SUNYSB.EDU 516-444-3126 *The opinions expressed above are my own and are not necessarily shared by the Microscopy Center*
Post-Doctoral Position in Analytical Electron Microscopy
The Structural Materials Research Section at the Pacific Northwest National Laboratory (PNNL) has an opening in the area of analytical electron microscopy. This is a one-year post doctoral position with the possibility of an extension for a second year. The research will involve high resolution compositional measurements at grain boundaries and interfaces as well as some microstructural analysis. Materials are primarily Al-based alloys and the work would support programs in interfacial deformation, recrystallization and stress corrosion cracking. The candidate is expected to have a Ph.D. in materials science, physics or a related discipline and expertise in utilizing EDS and/or PEELS with a transmission electron microscope for quantitative compositional analysis. Expertise in operation of a FEG-TEM is also beneficial.
PNNL is a national laboratory located on the Columbia River in SE Washington state. We have a well-equipped microscopy lab featuring a JEOL 2010F (Oxford EDS system and Gatan PEELS) in addition to several other microscopes (JEOL 1200, Philips 400, VG HB501) and associated sample preparation facilities.
For consideration, please submit a resume (including references) and publication list by Aug. 1, 1997 to:
Dr. John Vetrano Pacific Northwest National Laboratory MSIN P8-16 P.O. Box 999 Richland, WA 99352
When an instrument is operating the transformers and rectifiers in the HT=
tank generate heat, Whilst they are warming up the high voltage drifts t= o a level that is visible as a focus change on the screen over a few minute= s at 30,000X plus, or on a photograph at 15,000X with a 30secs plus exposur= e. Give the tank time to warm up such that there is thermal eqilibrium and=
the stability will or should be very good. One to one and a half hours should be sufficient in a SEM depending on kV and model. ALL electron optical instruments suffer from this problem its physics!
This is a bigger problem with high resolution TEM where a 100kV high voltage tank may take up to two hours to stabilise. A freon, or similar media, filled HT tank warms, even with a 100kV machine to give stability = in about 45 minutes!
Most people confuse focus drift with a final lens problem in the SEM wher= e as it is more likely high voltage change resulting in focus change.
} I need a lens with a "C"-mount for a Sony Model XC-77 analog video camera. } Can anyone recommend a source? A macro lens would probably work the best } for my application (image analysis). } } Thank you.
Hi Delilah,
What I have done is fit an adaptor mount to the video camera which enables me to attach an ordinary 35mm-camera lens. The brand of adaptor I have is Rowi ('cos that's all that was available at the time) but there are probably other manufacturers. The Rowi works fine though.
In my case I have attached a Nikon 50mm macro lens to do image analysis on large histological sections which are on a light box about 2 feet away. The adaptor works fine in these circumstances, but because the lens is not mounted directly on the camera (i.e. is separated from it by the adaptor "tube") it is not possible to focus on objects far away. In other words it is optically like fitting bellows or extension tubes between a camera and its lens.
In case you don't know what an adaptor is, it is simply a black tube; one end screws into the C-mount, the other end has a bayonet fitting as found on advanced 35mm cameras to which a 35mm camera lens can be attached - in my case it is a Nikon-type bayonet mount. Any large photo or video store should be able to help you.
Regards
Stephen Edgar
Electron Microscope Unit, Pathology Department School of Medicine University of Auckland Private Bag 92019 Auckland New Zealand
I agree with the previous posts regarding the supposed "safe limits" as being too high. I too have become sensitized from years of exposure to low levels. If I catch even a whiff of glut. my throat becomes irritated and I cough for a couple of days. I insist that it always be dealt with in a fume hood, no exceptions. Just my two cents worth.
cheers ed
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
Related to the cryo ladel and safety thread, does anyone know of a source for a dewar with a pouring spout?
We keep our LN2 in either a 5L or 10L dewar. Expecially for the SEM EDAX system, trying to pour from the 5L into the detector's vessel up near the ceiling is a backbreaking job. So we are left filling a very small (500 ml?) dewar about a million times going back and forth to finally fill the detector. As if this wasn't tedious enough even the small dewar doesn't pour well. (We've avoided using plastics such as polypropylene tripour beakers after having an exploding funnel episode of our own.) What would be great would be a ~1L dewar, kind of shortish, with a pouring spout. Anyone know of such a beast??
Thanks, Karen
-- Karen Zaruba kszaruba-at-mmm.com 3M Company, 3M Center Bldg. 270-1S-01 (612)737-2971 St. Paul, MN 55144 fax: 736-1519 "The opinions stated above are my own, not necessarily 3M's"
} } I need a lens with a "C"-mount for a Sony Model XC-77 analog video camera. } } What I have done is fit an adaptor mount to the video camera which } enables me to attach an ordinary 35mm-camera lens. snip } In my case I have attached a Nikon 50mm macro lens to do image } analysis on large histological sections which are on a light box } about 2 feet away.
I use the same setup on an XC-77 and it works very well. If you have no local supplier for the adapter, you can contact Edmund Scientific. They have a website and a (US) 800 number.
Let me add my voice-in-the-wilderness to this issue. I'm slightly sensitive to glut, not so bad I can't use it, but gloves and fume hoods are a *must*. Anyone not sensitized should demand them to prevent getting sensitized. Same for formaldehyde.
The problem doesn't end with these compounds, though--I've become sensitized to cacodylate and MS-222 (an anesthetic), and others have remarked on becoming sensitized to embedding resins. I would think the chronic exposure limits are too high generally, but in all cases gloves and fume hoods should be used more than they are.
The question is, what is to be done with anatomy classes? I've taken to recommending that any women who are pregant or working on it to seriously consider dropping the course.
Phil
P.S. There was a thread awhile ago on what gloves to use with what chemicals--was there a definitive list that came out of that discussion? I know glut goes through latex like the glove isn't there. P
} Sic Hoc Legere Scis Nimium Eruditionis Habes { 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 ******************
Most BSE detectors are said to be able to resolve about a tenth of an atomic number, particularly at the light element end of the spectrum. Ad= d a multi channel analiser and even better performance is possible.
As the accelerating voltage is decreased the efficiency of the detector will decrease due to detector conversion efficiency dropping along with specimen signal levels.
If the detector is from the 80s one would not expect to obtain results fr= om an accelerating voltage of much less than 15kV with a WD of 15 to 20 mm (detector specimen distance of 10 to 15mm).
If the detector comes from the early 90s we would expect to be able to wo= rk down to 5kV provided the electron gun was well adjusted giving us a good level of probe current.
This is for the jury-riggers out there and is definitely not UL or OSHA approved, but I think it is less hazard prone than a lot of LN2 ladling and pouring.
I have a 20 liter dewar for which I built a simple system to transfer LN2 at floor level to an EDS dewar at a height 63 inches.
Total cost is about 10 to 20 dollars plus a few hours of assembly. It requires a source of low pressure air.
The system uses a 6 foot length of 3/8" copper tubing insulated with dense foam tube, some PVC pipe, foam insulation spray, air couplings, two O-rings and a teflon seat ball valve.
My dewar plug is made from 3 inches of PVC pipe of the same diameter as the dewar throat. I machined some 'O' ring seats along its outside for a snug fit and seal. The long outlet tube and short air inlet tube are placed through the pipe and set in place with household foam insulation in a spray can. You may need to make an armature to hold the tubing in place during the cure. Try not to remember me while you are messing with this wet sub-foam slop. The inlet tube and outlet tube should not be in contact. The outlet tube should extend to near the bottom of the dewar and the inlet tube should be cut off at the bottom of the plug. The valve is on the inlet supply side.
I hold the plug while dispensing. If high pressure air is used and no restraint applied to the plug it could be blown out of the dewar neck. This and careless overfill are the only dangers unique to the apparatus I can think of.
A simpler system could probably be built with a two holed rubber stopper and a bracket to secure it in the dewar's throat. Stopper rubber, however, might not hold the copper tubing which immediately drops to LN2 temps and would develop a lubricating layer of ice and water. Keeper sleeves could be soldered onto the copper tubing on the underside of the stopper.
Transfer is by pressurizing the sealed dewar through the valved inlet tube with about 10 pounds of compressed air. I safely transfer 7.5 liters of LN2 in about 30 seconds with no back strain or sight of the liquid. If the exit tube is just the right length one can drain the dewar to the last "drop" without tipping the vessel.
A styrofoam float with a calibrated stick long enough to poke through the dewar's throat can be used to monitor fill progress when the destination dewar is located up high.
This is for the jury-riggers out there and is definitely not UL or OSHA approved, but I think it is less hazard prone than a lot of LN2 ladling and pouring.
I have a 20 liter dewar for which I built a simple system to transfer LN2 at floor level to an EDS dewar at a height 63 inches.
Total cost is about 10 to 20 dollars plus a few hours of assembly. It requires a source of low pressure air.
The system uses a 6 foot length of 3/8" copper tubing insulated with dense foam tube, some PVC pipe, foam insulation spray, air couplings, two O-rings and a teflon seat ball valve.
My dewar plug is made from 3 inches of PVC pipe of the same diameter as the dewar throat. I machined some 'O' ring seats along its outside for a snug fit and seal. The long outlet tube and short air inlet tube are placed through the pipe and set in place with household foam insulation in a spray can. You may need to make an armature to hold the tubing in place during the cure. Try not to remember me while you are messing with this wet sub-foam slop. The inlet tube and outlet tube should not be in contact. The outlet tube should extend to near the bottom of the dewar and the inlet tube should be cut off at the bottom of the plug. The valve is on the inlet supply side.
I hold the plug while dispensing. If high pressure air is used and no restraint applied to the plug it could be blown out of the dewar neck. This and careless overfill are the only dangers unique to the apparatus I can think of.
A simpler system could probably be built with a two holed rubber stopper and a bracket to secure it in the dewar's throat. Stopper rubber, however, might not hold the copper tubing which immediately drops to LN2 temps and would develop a lubricating layer of ice and water. Keeper sleeves could be soldered onto the copper tubing on the underside of the stopper.
Transfer is by pressurizing the sealed dewar through the valved inlet tube with about 10 pounds of compressed air. I safely transfer 7.5 liters of LN2 in about 30 seconds with no back strain or sight of the liquid. If the exit tube is just the right length one can drain the dewar to the last "drop" without tipping the vessel.
A styrofoam float with a calibrated stick long enough to poke through the dewar's throat can be used to monitor fill progress when the destination dewar is located up high.
I am looking for some direction on the preparation of diverse protozoans (several common ciliates, Euglena and A. proteus) for scanning electron microscopy. My major difficulties lie in the areas of keeping them "relaxed" during fixation and getting them to adhere well to some substrate such as polycarbonate filters, PLL coated coverslips, etc. Additionally, fixation of the Amoeba is a disaster for me, although the others look OK with a glut/osmium mix.
Any tips or references would be greatly appreciated; thank you in advance.
Dick Briggs Biology Department Smith College Northampton, MA 01063
I want to do some image processing and analysis of some secondary electron images of mineral grains. These are from grain mounts rather than polished mounts and are available as 8 bit grayscale images. The grains are not separated nicely against background but rather are piled up, giving substantial overlap. However, a sufficient number of grains are not overlapped that some assessment of their properties should be possible. I am able to segement the images to a binary state (all pixels 0 or 255) but attempts to find and analyse the objects are not very successful because partially hidden grains fade into the dark background and do not segment cleanly.
Does anyone have experience of doing this kind of processing? Is it possible? If anyone can give me a list of steps to follow I'd be most grateful.
NB I'm doing the image processing/analysis in Image-Tool 1.27.
If you choose to pressurize your LN2 dewar, be careful of condensing Liq Oxygen in it. The oxygen in the pressurizing air condenses at a higher temperature than the LN2 so that it is possible to wind up with a large amount of Liquid oxygen in the dewar. Liquid oxygen is hazardous to deal with. This is particularly a problem if you never completely the dewar as the LOX can build up over time. Pressurizing with N2 gas is MUCH safer.
Henk Colijn
} } This is for the jury-riggers out there and is definitely not UL or OSHA } approved, but I think it is less hazard prone than a lot of LN2 ladling } and pouring. } } I have a 20 liter dewar for which I built a simple system to transfer } LN2 at floor level to an EDS dewar at a height 63 inches. } } Total cost is about 10 to 20 dollars plus a few hours of assembly. It } requires a source of low pressure air. } {snip}
--- On Thu, 05 Jun 1997 15:48:24 -0400 "Pitzenberger, Marcia H." {marcia_pitzenberger-at-merck.com} wrote:
} I'm getting some human hepatocytes for possible DAB staining within the } next couple weeks. I have no experience with human hepatocytes or DAB. } Can anyone recommend an appropriate fixative and buffer? Also, how long } can I store these and they'll be o.k. for DAB staining? Should I store } them in buffer after a period of fixation? Any help / advice would be } appreciated. } } Marcia Pitzenberger } Merck & Co., Inc. } P.O.Box 4, WP45-251 } West Point, Pa 19286-0004 } (215)652-9767 } marcia_pitzenberger-at-merck.com } -----------------End of Original Message-----------------
Dear Marcia,
In our lab we have routine protocols for DAB staining of human neutrophils, so you may have to adapt a little.
Fixation: 1 hour at 4-6°C (on ice) with 3% glutaraldehyde in 0.1M cacodylate buffer, pH 7.35.
They are washed 3x and stored overnight in 0.1M cacodylate 7% sucrose buffer, pH 7.35.
The sooner DAB staining occurs the better. Activity falls off such we try to have the samples stained within 2 weeks (3 at the outside). The hepatocytes may differ from that however.
The DAB staining that we do is for myeloperoxidase (0.01% H2O2 for 30 min at room temp in the dark) but it also works for the various catalase procedures. If you require any references, I may be able to provide some. Let me know.
Hope this helps,
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
Thanks for your suggestion. Although your method may be well known in the stereological literature, it probably isn't to microscopists. It would be nice for you to furnish an appropriate citation or two.
On Thu, 5 Jun 1997 DrJohnRuss-at-aol.com wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } In a message dated 6/5/97 2:02:29 PM, you wrote: } } } On Wed, 4 Jun 1997, Corazon D. Bucana wrote: } } } } } I am trying to compare the distribution of particles in histological } tissues } } } and I would like to determine the distance(s) of particles relative to } their } } } nearest neighbor. Since the particles are not of uniform size I thought } } } that if I determined the centroids it will give me a reproducible } parameter. } } If the distance you really want is the separation between centroids of the } sections in the plane, you can get if from their centroids. If what you } really want is the 3D distance between the surfaces of the particles, you can } get it by drawing random lines (which are random in 3D if your section was } random) on the image and measuring the length of the intercept lines across } the inter-particle background. This is a tad complicated sounding, but } actually rather simple. Bear with me } Each intercept length we will call L. You want to determine the average value } of the reciprocal, 1/L. } The average value of 1/L is two-thirds of the value of 1/T where T is the } actual mean interparticle spacing (surface to surface). } The proof is geometrical and somewhat arcane, but well known [;-)] in the } stereological literature. } } John Russ } }
Edmund Glaser, D. Eng. Dept. Physiol. Univ. Md. School. Med. Baltimore, MD 21201 USA Ph: (410) 706-5041 Fax: (410) 706-8341
I have a customer half way across the country who would like to examine the possibly corroded surface of a large mold in a plastics-forming plant. The mold is too big to be put in the SEM and it cannot be sectioned (big $). I suggested making a plastic film replica of the surface and sending it to me.
I can't seem to find my references/instructions for making acetate (?) replicas with acetone and I want the customer to be able to do it right (we have only one chance).
Any guidance would be appreciated, even from vendors. :-)
------------------------------------------------ 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 --
Advice needed, We are about to purchase a high vac metal evaporator for shadowing and carbon deposition, and would like some recommendations. We need a dependable workhorse, as this equipment will be destined for many core users. High vac (10 -7 mbar) within a reasonable time, durability and ease of use is what we want. We are currently looking at the Edwards Auto 306 (possibly with a cryo-pump, more likely with a diff pump) and Denton's DV-502A (we have between 15-20K to spend). We have been using an old Polaron which is on its last legs. All thoughts and suggesstions are greatly appreciated.
Mike Delannoy JHMI Microscopy Facility (410) 955-1365
In a message dated 6/6/97 10:04:09 AM, eglaser-at-umabnet.ab.umd.edu wrote:
} Thanks for your suggestion. Although your method may be well known in } the stereological literature, it probably isn't to microscopists. It } would be nice for you to furnish an appropriate citation or two.
H. J. Gundersen et al, 1978, J. Microscopy v.113, p.27 J. C. Russ, 1986, Practical Stereology, Plenum, New York, p. 62
I can't resist the opportunity to point out that microscopists SHOULD study the stereological literature - they are examining 2D images of sections cut through 3D structures, and if they ignore the stereological meaning of their 2D measurements (no matter how carefully done) the results will not be meaningful. End of soapbox.
Dear Roger, here at Noesis Vision Inc we have lots of experience with particle separation. We have powerful separation algorithms in Visilog, send us your images and we can do a feasibility study for you at no charge.
Regards,
At 08:41 AM 6/6/97 -0230, Roger Mason wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
---------------------------------------------------------------------------- ------------- Luc Nocente Tel: 514 345 1400 Noesis Vision Inc. Fax: 514 345 1575 e-mail: ln-at-noesisvision.com http://www.noesisvision.com 6800 Cote de Liesse, Suite 200 St-Laurent, PQ H4T 2A7,Canada ---------------------------------------------------------------------------- -------------
We use old metal coffee pots for LN2. They have straight sides, have a pour spout, and plastic insulated handles. Well-designed to keep hot coffee from your hands, they do good service with liquid nitrogen. Water will condense on the outside and freeze, but this doesn't get into your dewars if transit time is reasonable. They are aluminum and available in most hardware stores in various sizes from 0.5 to 2 or more liters for around $10. They hang up well and don't break on falling or cooling.
I also like metal soup ladles, also from the hardware store, with about half liter capacity. Cost about $4. These are spot welded and come apart after a few years of jolting about. We keep half dozen hanging over our work area so that they warm up and dry out between uses.
Jacob
Jacob Bastacky, M.D. Room 116 Donner Laboratory Lawrence Berkeley Laboratory EMail: sjbastacky-at-lbl.gov University of California Phone: (510) 486-4606 Berkeley, California 94720 Fax: (510) 486-4750
Does anyone have suggestions of software for analysing negatives taken on a TEM with negatively stained material? (I'm currently using NIH-Image, but would like to render, if possible, 3D volumes of my material)
Also, I wonder if there's any known way to estimate/calculate the height of structures from negatively stained material scans...
In message {3396DE9A.5AF-at-accessone.com} , Bart Cannon {cannonmp-at-accessone.com} writes } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
What about a source of low pressure nitrogen - like for instance a dewar of LN2? (g)
Drop a small heating coil in the dewar. -- Anthony James Bentley Surface Data Scientific Instrumentation and Software Web site http:\\www.surface.demon.co.uk
I must confess I hadn't thought about oxygen condensation.
Luckily my design uses no sparkable materials.
While on the subject I wonder how my system would produce more gas stratification that any sitting, capped dewar since the leak tight cap is only used for 30 seconds.
This inspires the question: are all capped dewars oxygen bombs???
Some LN2 can be transferred from the introduction of the transfer tube into the LN2, but not enough for a fill. The compressed air is so quick and easy I hadn't bothered to look into N2 as a pressurizing gas.
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Hi All-
I'm looking to purchase a sample holder to use with our stereo microscope. I need it to hold samples from one eighth inch to several inches and to be adjustable (turn the sample). I've looked through all of our catalogues with no success. Any ideas?
All over our building we have an argument raging as to the correct formulation for PBS. Mainly the argument consists of opinions that the phosphate concentration should be 0.1M, and other opinions that the concentration ought to be O.OlM. (We all agree that NaCl concentration is 0.15M). There are also conflicting statements in the literature as to the composition of PBS. What we need is a thorough explanation of why one or the other concentration of phosphate salts is correct for mammalian tissue for TEM in procedures such as the DAB process. I would so much appreciate help in logically settling this argument around here. So long, Hildy
Hildy Crowley University of Denver 2101 E. Wesley Ave Denver, CO 80208
Replicas are easy to make. Cut the Acetate to size then place a drop or two of acetone on surface to be replicated. Place the acetate on surface. The acetone will soften the acetate so you will want to apply slight pressure to ensure acetate molds to surface. I use a cotton swap, but have also used my finger.
If you need to do the top surface put the acetone on the acetate.
You can by a kit from Bueller (most likely spelled wrong) that has foil on the back which allow viewing with an optical microscope.
The first replica will clean the surface, save it for analysis as in this case it will contain corrosion products.
Do more than one just incase something goes wrong. (it does for me.)
For those of you who were looking for the Leaf 45, I think that I found the site for the new improved version of it. Check this site out http://www.hyperzine.com/photokina/brem.html
Here is a short paragraph from the page.
Bremson Negative Scanner
photokinaBremson Inc. announces the debut of the versatile new Bremson 45HS Scanner, a high resolution, multi-format negative scanner designed by Leaf Inc., for exclusive manufacture and distribution by Bremson Inc. Well below the price of comparable high-end, high cost drum scanner technology, the affordability of the new 45HS Scanner is a welcome addition to commercial and other labs that specialize in digitizing negatives.
I have no interest in this company and was surfing the net looking for negative scanners. No one seemed to know precisely what happened to the Leaf 45.
If you are looking into scanners, here's a place to start.
This is a site by the Scientific Photography Lab at the University of Basel that I found that has a long list of scanners and information on them separated by formats:
http://www.foto.unibas.ch/scanners.html
Very informative if you are looking for a scanner.
Richard, I was reading my wife's e-mail and noticed your question. I am an occupational hygiene chemist and have been measuring glutaraldehyde levels in hospitals and clinics throughout Queensland for about six years, and I agree that 0.2ppm is far too high. Following representations from a number of people, myself included, Worksafe Australia have reduced the limit to 0.1ppm . I dont consider myself to be particularly sensitive to glutaraldehyde, but I know that 30mins exposure to 0.09ppm will guarantee a headache. In my experience, to avoid complaints from staff, the level should be kept below 0.03ppm if possible, although I did come across someone who became sensitised at 0.02ppm while working in an X-ray dark room. X-ray film fixer contains a small amount of glutaraldehyde some of which gets blown around the room when the film is dried. I usually use a Glutaraldemeter for the measurements, and it is quite good unless there is alcohol in the room. However, it is so obvious when the meter starts to wind off scale, that it is not a problem.
The Glutaraldemeter is good in that it gives spot measurements, other methods I have used such as 2,4DNPH tubes give an average reading over time, but are not so good where glutaraldehyde is being used sporadically.
Imust say that I havent come across an EM lab without a fume hood, and so they dont seem to have your problem.
Hope this helps.
George Lee Chemist- Occupational Hygiene Queensland Health Scientific Services PO Box 594 Archerfield 4108 Brisbane Queensland
Edmund Glaser wrote: } } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Thanks for your suggestion. Although your method may be well known in } the stereological literature, it probably isn't to microscopists. It } would be nice for you to furnish an appropriate citation or two. } } On Thu, 5 } Jun 1997 DrJohnRuss-at-aol.com wrote: } } } ------------------------------------------------------------------------ } } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } } -----------------------------------------------------------------------. } } } } } } In a message dated 6/5/97 2:02:29 PM, you wrote: } } } } } On Wed, 4 Jun 1997, Corazon D. Bucana wrote: } } } } } } } I am trying to compare the distribution of particles in histological } } tissues } } } } and I would like to determine the distance(s) of particles relative to } } their } } } } nearest neighbor. Since the particles are not of uniform size I thought } } } } that if I determined the centroids it will give me a reproducible } } parameter. } } } } If the distance you really want is the separation between centroids of the } } sections in the plane, you can get if from their centroids. If what you } } really want is the 3D distance between the surfaces of the particles, you can } } get it by drawing random lines (which are random in 3D if your section was } } random) on the image and measuring the length of the intercept lines across } } the inter-particle background. This is a tad complicated sounding, but } } actually rather simple. Bear with me } } Each intercept length we will call L. You want to determine the average value } } of the reciprocal, 1/L. } } The average value of 1/L is two-thirds of the value of 1/T where T is the } } actual mean interparticle spacing (surface to surface). } } The proof is geometrical and somewhat arcane, but well known [;-)] in the } } stereological literature. } } } } John Russ } } } } } } Edmund Glaser, D. Eng. } Dept. Physiol. } Univ. Md. School. Med. } Baltimore, MD 21201 USA } Ph: (410) 706-5041 } Fax: (410) 706-8341
I never subscribed to this bloody list so please stop all this friggen mail from comming here...
Harold, plastic replicas for SEM are generally easy to prepare. Preparation of a second stage, perhaps in Pt/C is much more tricky, but not needed for SEM. I used Bioden acetate film for many years but any thin plastic replicating material should do well. Bioden is used with acetone or methyl acetate.
Cut a piece of film material to size. Place on top- of the region for replication. Drop solvent onto plastic until its just wet all over. Apply gentle pressure to the film just after solvent has just evaporated to ensure good contact. Wait for at least 10 minutes before pulling replica off. Pull replica off and throw this into the bin. Repeat and "bin" second replica - to clean surface. Retain third and subsequent replica for use. Metallise (sputter) specimen surface before mounting and viewing.
Notes: Handle edge of replica and only with tweezers. Mark replica's specimen site like sheet film notching; when facing (emulsion or replica) cut the top right corner. When viewing in SEM reverse the polarity to gain a realistic "hills and valleys" impression. Single stage replicas are reversed. You could look at a "bin" replica, after carbon coating in backscattered mode to check any of the extracted material and also to do EDS on that. Angle shadow casting with Pt (from wire wrapped around a tungsten 'V' filament) and subsequent carbon coating sometimes enhances features. 10 to 30 degree angles of evaporation are used. Low angles are best for replicas with very little topography. 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 } } I have a customer half way across the country who would like to examine } the possibly corroded surface of a large mold in a plastics-forming } plant. The mold is too big to be put in the SEM and it cannot be } sectioned (big $). I suggested making a plastic film replica of the } surface and sending it to me. } } I can't seem to find my references/instructions for making acetate (?) } replicas with acetone and I want the customer to be able to do it right } (we have only one chance). } } Any guidance would be appreciated, even from vendors. :-) } } ------------------------------------------------ } 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} }
Hildy: What is the DAB process? The primary function of a buffer is to set and hold the pH at the desired level. 0.1M of the phosphates is normally used with mammalian tissues and the addition of salts to a fixative is usually not required. I remember that NaCl is 8.5g/litre in tissue fluids - and that translates almost exactly to your 0.15M figure. 0.1M buffer with the addition of that salt is certainly hypertonic. Soerenson developed the phosphate based buffer recipe. The addition of 8.5g NaCl (or a more complex balanced salt solution) turns that into PBS. No doubt, people have different fancies (experiences, data?) as to what strength the buffer should be. I suggest that 0.01 is low and 0.1M is on the high side for PBS. 0.01M is about the minimum effective buffer concentration for fixatives. That level without salt is commonly used for plants since they have much lower osmolarity than mammals do. Now, had I known what the DAB process is, I might have kept quiet! 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 } Dear Folks, } } All over our building we have an argument raging as to the correct } formulation for PBS. Mainly the argument consists of opinions that the } phosphate concentration should be 0.1M, and other opinions that the } concentration ought to be O.OlM. (We all agree that NaCl concentration } is 0.15M). There are also conflicting statements in the literature as to
} the composition of PBS. } What we need is a thorough explanation of why one or the other } concentration of phosphate salts is correct for mammalian tissue for TEM } in procedures such as the DAB process. } I would so much appreciate help in logically settling this argument around } here. } So long, } Hildy } } Hildy Crowley } University of Denver } 2101 E. Wesley Ave } Denver, CO 80208 }
Harold Crossman has raised some questions about the use of "acetate" replicas for SEM. The material is actually cellulose acetate, and it is available from several sources, including my employer, Structure Probe/SPI Supplies :-)
The use this material goes back to the days when fractography was done by TEM because SEM was not commercially available. There is, therefore, some very old literature on how to use the material, but since most of it is art, anyway, most current authors assume that the reader is generally familiar with the technique.
There are actually two purposes in using this material. Even for relatively small samples, it may be necessary to remove corrosion products in a nondestructive manner, and in my opinion the best way to do this is to use acetone-softened cellulose acetate, which will remove the corrosion product but leave what is left of the metal. I know about various "nondestructive" chemical removal agents, and I have done enough testing of these agents that I question whether they are truly nondestructive. As Harold points out, we have one chance at a "real" failure.
The second purpose is to make a replica of the cleaned fracture surface for examination in the microscope. Whichever the purpose, the technique is basically the same.
For TEM replication, historic practice has generally been to dissolve the replicating material in a suitable solvent at a concentration around 2%, put a drop on the area of interest, place a grid on the drop and go away for a while. Replicating materials with which I have some familiarity include collodion, parlodion, formvar, butvar, pioloform, poly(acrylic acid) and good old cellulose acetate; some of these material names are trademarks and should be capitalized; there is an appropriate solvent for each, and part of the "art" is selecting the proper combination. For example, water, used to dissolve poly(acrylic acid) is very suitable for some polymer surfaces but not suitable for most metals. When the solvent is completely evaporated, the grid can be picked up using tweezers, shadowed (or coated for SEM) and examined. This can work very well for TEM on the large, flat surfaces of those rare fatigue failures which wind up in textbooks, and it gives an unequivocal indication which part of the fracture surface generated which replica, but it has problems for "real" fractures in the SEM.
The basic problem is that the procedure is too good. The solution gets into every crack and crevice of the fracture surface, and the replica is "locked" to the surface so that it is impossible to remove without tearing. For practical SEM use, then, normal practice has been to use "tape".
Cellulose acetate is available from various suppliers (including SPI Supplies) as tape, a strip of material about 2.5 cm wide, from which the piece to be used is cut. Also available from some suppliers are sheet product. There tend to be different thickness preferences; all thicknesses work, but some folks like it thicker, and others like it thinner. In use, the area to be replicated is moistened with the solvent (acetone for cellulose acetate), the tape is also moistened on the side that will form the replica, and the solvent is allowed to soften the tape enough, but not too much (yes, there is some "art" here). I find that forming a shallow "cup" in the tape helps me to hold the solvent in the area of interest without making a mess. The obvious idea of dipping the tape into the solvent is a recipe for a real mess, because the next step will fail.
When the tape has softened enough, but not too much, it is carefully pressed into the drop of solvent on the surface of interest. Thorough contact of wetted surfaces is essential. Our "official" tool for pressing is the eraser on a pencil, but when the situation is serious, I use my right thumb; I suppose that my left thumb would work, but I've never tried :-) If the tape is too soft, the pressing tool will go through the tape, ruining the replica; if the tape is not soft enough, it will sit above the surface of interest and not capture its fine features. This is an art, not an "exact" science.
The next step is the most critical--go away for a while. Most "bad" replicas were destroyed by being "pulled" too quickly; it is essential that the solvent be completely evaporated, or the fine features will be left on the fracture surface. When completely dry, the replica is removed by pulling the tape upward. With the obvious exception that the tape may be attacked by the solvents in various "paint" products, the replica can be prepared like any other nonconductive SEM sample and examined at a remote location.
In actual practice, I do ten times as much replication to clean fracture surfaces as I do to examine them. The "first pull" replica from cleaning the fracture surface provides a "clean" sample of the corrosion product without any interference from the substrate, and in some cases, this is actually a better procedure than attempting to analyze a corrosion product still on the substrate.
Two disclaimers, first, SPI Supplies has an obvious interest in promoting the use of replicating products, including cellulose acetate tape, and second, replication involves the use of volatile solvents, some of which are somewhat "nasty". My first experience with TEM was making replicas using collodion dissolved in amyl acetate; not knowing anything about amyl acetate at that time, I can only observe that I developed the habit of driving 90 miles an hour on the way home after breating the stuff all day. Please be careful.
I would be happy to try to answer specific questions from my experience. The problem Harold presents is a very real one--he has only one chance to get the replicas, using untrained staff, and good replication is an art which requires experience.
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
Has anyone heard about a reference called "Ranking of the cathodes of Round-Robin tests 1 and 2 according to the structure data". Jernkontoret 1988.? I don't know if it is a book or a paper, neither where to find it.
In the "old" pre OSHA days we used a 10 watt resistor taped to the bottom of the transfer tube with a resistance that allowed it to be wired directly to 115 VAC. This could be plugged in for long enough to build up the pressure needed for transfer. Today the 115 VAC would be considered too hazardous.
However there is a way to make everyone happy. Get one of the many 1 amp, 12 volt in-line or wall mount power modules that are available (115 VAC in and 12 VDC 1 A out). You can use the same principal as above but now with an intrinsically safe voltage. No need for any pressurizing gas at all.
I have been offered a donation of a working Zeiss EM-6 SEM by someone who doesn't know anything more about it than its name. Can anyone tell me anything about this scope?(e.g., age, size, reliability, parts availability, resolution, ease of operation)
Is there anyone else out there who could use this donation?
thanks
steve
----------------------------------------------------------------------------- Dr. Steven Barlow Electron Microscope Facility Dept. of Biology San Diego State University 5500 Campanile Drive San Diego CA 92182-4614 phone: (619) 594-4523 fax: (619) 594-5676 email: sbarlow-at-sunstroke.sdsu.edu http://www.sci.sdsu.edu/EM_Facility
One solution that we use is a set of steps. Our carpenter shop built (over built!) a set of wooden stairs, three steps high, that sits behind our SEM. We just climb the stairs with a 4 liter dewar when we need to fill the EDS dewar. That way we are about chest high with the opening of the EDS dewar and pouring is easy. It also prevents any LN2 from splashing down on us while filling. You can buy portable metal stairs in catalogs such as Grainger.
By the way we have 12 foot high ceilings in the SEM room!
Louie Kerr
At 3:46 PM -0500 6/5/97, kszaruba-at-MMM.COM wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Louie Kerr Research and Education Support Coordinator Marine Biological Laboratory 7 MBL Street Woods Hole, MA 02543 508-289-7273 508-540-6902 (FAX)
Apologies to the list, but I've lost their email address, and can't find their posts in the archives.
Mizuho Shimizu from this company recently posted about a Nipkow disc confocal microscope.
Would someone who kept their address please send me their email address? Or if this person, or someone from their company is reading this, please contact me.
Thank you.
Phil
} Sic Hoc Legere Scis Nimium Eruditionis Habes { 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 ******************
How much would you think, might be the value of a light microscope with no own light source, several objectives (max. 60 x) and brass exterieur ? My great-grandfather as a passionate precision mechanic built it himself around 1910, and now somebody would like to buy it, but we have no idea how much is a sensible price.
} Date: Mon, 9 Jun 1997 09:54:07 -0500 } To: HILDEGARD CROWLEY {hcrowley-at-du.edu} } From: Tom Phillips {tphillips-at-biosci.mbp.missouri.edu} } Subject: Re: TEM:Help!Correct PBS formula???? } Cc: } Bcc: } X-Attachments: } "Saline" is a physiological term for 0.9% NaCl solution which is 0.154 M. This is osmotically correct for human serum. If you add a large amount of phosphate buffer (e.g., 0.1M), one would have to lower the NaCl concentration in an equivalent manner if you want to maintain osmolarity. The Gibco catalog gives two formulations for PBS on page 2.9 of their 1997 catalog. one lists 9.0 g NaCl, 0.795 g NaH2POH-7H2O and 0.144 g KH2PO4 (all per liter). this is about 3 mM Na2PO4 and 1 mM KH2PO4. the other formulation is 0.21 g KH2PO4 and 0.726 g NaH2PO4-7H20. either one causes a slight hyperosmolarity in respect to straight "saline". In osmicated tissue, osmolarity might not be important but I suspect in aldehyde fixed tissue it can be. In TEM, one has to worry about PO4 based buffers causing precipitation of Ca salts. In procedures like DAB, one has to ensure the amount of buffer capacity is not exceeded. } } } All over our building we have an argument raging as to the correct } } formulation for PBS. Mainly the argument consists of opinions that the } } phosphate concentration should be 0.1M, and other opinions that the } } concentration ought to be O.OlM. (We all agree that NaCl concentration } } is 0.15M). There are also conflicting statements in the literature as to } } the composition of PBS. } } What we need is a thorough explanation of why one or the other } } concentration of phosphate salts is correct for mammalian tissue for TEM } } in procedures such as the DAB process. } } I would so much appreciate help in logically settling this argument around } } here. } } So long, } } Hildy } } } } Hildy Crowley } } University of Denver } } 2101 E. Wesley Ave } } Denver, CO 80208 }
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)
Biological Physics. Postdoctoral position at the University of Virginia. A postdoctoral position is available in the Department of Molecular Physiology and Biological Physics of the School of Medicine. The research projects in which the applicant is expected to play a major role involves electron energy loss spectroscopy and energy filtered scanning transmission electron microscopy. The position will include both development of software and instrumentation for achieving 2 to 3 nm spatial resolution compositional imaging and hands-on application of the method to significant biological problems. The laboratory has been engaged in NIH- supported research developing and applying analytical electron microscopy for 25 years through an interdisciplinary program based on the collaboration between physicists and biologists. Equipment available includes a 200kV electron microscope equipped with field emission gun (Philips CM200-FEG), a GATAN electron spectrometer adapted to our own CCD camera, a CM12 electron microscope and two energy dispersive X-ray detectors. Investigators in the program are also members of the Center for Structural Biology of the University of Virginia and have programs involving collaborations with the X-ray crystallography and atomic force microscopy groups and with molecular biologists and investigators engaged in other biological disciplines.
Candidates should have a Ph.D. in Physics, material science or engineering and be familiar with computer programming, instrumentation and, preferably, experience with electron microscopy and electron energy loss spectroscopy. Applications with biographical sketch, bibliography and the names of three references should be sent to: Dr. Andrew P. Somlyo, Department of Molecular Physiology and Biological Physics, University of Virginia, P.O. Box 10011, Charlottesville, VA 22906-0011, USA. The University of Virginia is an Equal Opportunity/Affirmative Action Employer.
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Dear specialists,
How much would you think, might be the value of a light microscope with no own light source, several objectives (max. 60 x) and brass exterieur ? My great-grandfather as a passionate precision mechanic built it himself around 1910, and now somebody would like to buy it, but we have no idea how much is a sensible price.
I've never subscribed to this list so please stop all these mailings....
Imager wrote: } } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Edmund Glaser wrote: } } } } ------------------------------------------------------------------------ } } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } } -----------------------------------------------------------------------. } } } } Thanks for your suggestion. Although your method may be well known in } } the stereological literature, it probably isn't to microscopists. It } } would be nice for you to furnish an appropriate citation or two. } } } } On Thu, 5 } } Jun 1997 DrJohnRuss-at-aol.com wrote: } } } } } ------------------------------------------------------------------------ } } } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } } } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } } } -----------------------------------------------------------------------. } } } } } } } } } In a message dated 6/5/97 2:02:29 PM, you wrote: } } } } } } } On Wed, 4 Jun 1997, Corazon D. Bucana wrote: } } } } } } } } } I am trying to compare the distribution of particles in histological } } } tissues } } } } } and I would like to determine the distance(s) of particles relative to } } } their } } } } } nearest neighbor. Since the particles are not of uniform size I thought } } } } } that if I determined the centroids it will give me a reproducible } } } parameter. } } } } } } If the distance you really want is the separation between centroids of the } } } sections in the plane, you can get if from their centroids. If what you } } } really want is the 3D distance between the surfaces of the particles, you can } } } get it by drawing random lines (which are random in 3D if your section was } } } random) on the image and measuring the length of the intercept lines across } } } the inter-particle background. This is a tad complicated sounding, but } } } actually rather simple. Bear with me } } } Each intercept length we will call L. You want to determine the average value } } } of the reciprocal, 1/L. } } } The average value of 1/L is two-thirds of the value of 1/T where T is the } } } actual mean interparticle spacing (surface to surface). } } } The proof is geometrical and somewhat arcane, but well known [;-)] in the } } } stereological literature. } } } } } } John Russ } } } } } } } } } } Edmund Glaser, D. Eng. } } Dept. Physiol. } } Univ. Md. School. Med. } } Baltimore, MD 21201 USA } } Ph: (410) 706-5041 } } Fax: (410) 706-8341 } } I never subscribed to this bloody list so please stop all this friggen } mail from comming here...
As "News and Commentary" Editor for the journal "Microscopy and Microanalysis," I put a "Future Meetings" section in vol 3.2, p. 167, and "Short Courses" on p. 170. We allowed the input of extended announcements for events that were scheduled to occur close to the date of vol 3.2 (March into Fall) and brief notices for events occurring later (1998 onwards).
I plan to put in an updated Future Meetings and Short Courses section in vol 3.5, which has an input closing date to me of July 14 and will be mailed on September 24th.
I invite organizers of meetings and courses to send input for events occurring from October onwards. Extended listings will be considered for events in the October to February 1998 time period and shorter listings for later events. We will repeat short listings up to the date of the event but will allow only one extended listing. Submitters should plan accordingly.
Please send input via e-mail, offline. Please follow the format in vol 3.2, p. 167. Text submissions mailed to me will be typed by me on a time available basis. (Scary thought! :-) ) Send e-mail as plain ascii text. No attached word processor formatted documents please.
The journal goes to about 5,000 microscopists world-wide. International listings are welcomed.
Ron Anderson ron-anderson-at-vnet.ibm.com or, -at- IBM, Mail Stop E-40 Hopewell Jct., NY 12533 USA (see "scary thought," above)
This query was forwarded to me by a friend at Los Alamos who asked if I would post it to the Listserver. We would be very grateful if anyone can advise Joanna McKittrick at UCSD on this matter. Her e-mail address is jmckittrick-at-ucsd.edu
Many thanks in advance,
Gill Bond Dept Materials & Met. Eng. New Mexico Tech
---------- Forwarded message ----------
} I have a technical question. Do you know of any computational } way to generate microstructures? For example, if I have a 2 } phase material, know the average and SD of the grain size for } each phase and the volume %, is there a way to generate } an image???? Do you know anyone who does this? } } Also, do you know anyone doing combustion research here? } I am interested in thermodynamic programs, too. It seems } with the tremendous computational facilities at LANL there } should be a lot of work done in these areas. } } Last--what is your favorite drawing program? I have been } using McDraw (yuk) and am looking for something a little } more powerful. } } Thanks! } } Joanna } } ************************************** } Joanna McKittrick } Department of Applied Mechanics and } Engineering Sciences and } Materials Science Program } University of California at San Diego } 9500 Gilman Drive } La Jolla, CA 92093-0411 } } 619-534-5425 (phone) } 619-534-5698 (fax) } jmckittrick-at-ucsd.edu } ************************************* }
VOTING IN PROGRESS FOR IMMUNOCYTOCHEMISTRY NEWSGROUP sci.bio.immunocytochem
If you want to see a NEWSGROUP dedicated to immunohistochemistry, immunocytochemistry and other affinity labelling methods, then please do try to vote if you havn't already done so. The votetaker must collect 100 more YES than NO votes for the group to be made official. The closing date for ballots to reach the votetaker is 16th June. All you need to vote is a valid e-mail address.
The 2nd (and final) CFV, or CALL FOR VOTES was posted to "news.announce.newgroups" and "news.groups" on the 6th June. Voting instructions and the ballot can be found at the end of the CFV.
Some people are having trouble accessing the CFV or "Call For Votes" via the Usenet Newsgroups. The VOTETAKER, David Bostwick, will SEND YOU the CFV (with voting instructions and the ballot) if you e-mail him bostwick-at-cas.chemistry.gatech.edu
Alternatively, you can go the the FTP site: ftp://ftp.isc.org/pub/usenet/news.announce.newgroups/sci/sci.bio.im munocytochem where all the old "Request For Discussions" are stored, together with the CFV (right at the end!)
Remember, the closing date for votes to reach the votetaker is 16th June, so hurry! Please feel free to e-mail me with any questions you may have about the proposed group or voting.
Many thanks for your very practical and timely tips on making replicas. Andy Blackwood's and Jim Darley's responses were so detailed that they may have saved me a trip to Indiana! I forwarded all the messages to my customer who will be able to handle the project locally.
------------------------------------------------ Opinions or statements expressed herein, rational or otherwise, do not necessarily reflect those of my employer.
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I am having problems with wrinkles in Epon thin sections (gold interference colors)and would like some advice or information about getting rid of them. The specimen is ameba and has been grown on the surface of an Epon bullet coated with a collagen matrix. The ameba are then fixed, dehydrated and embedded within the Epon bullet. Early sections (the ones first off) are of interest since the ameba tend to attach to substrate.
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Biological Physics. Postdoctoral position at the University of Virginia. A postdoctoral position is available in the Department of Molecular Physiology and Biological Physics of the School of Medicine. The research projects in which the applicant is expected to play a major role involves electron energy loss spectroscopy and energy filtered scanning transmission electron microscopy. The position will include both development of software and instrumentation for achieving 2 to 3 nm spatial resolution compositional imaging and hands-on application of the method to significant biological problems. The laboratory has been engaged in NIH- supported research developing and applying analytical electron microscopy for 25 years through an interdisciplinary program based on the collaboration between physicists and biologists. Equipment available includes a 200kV electron microscope equipped with field emission gun (Philips CM200-FEG), a GATAN electron spectrometer adapted to our own CCD camera, a CM12 electron microscope and two energy dispersive X-ray detectors. Investigators in the program are also members of the Center for Structural Biology of the University of Virginia and have programs involving collaborations with the X-ray crystallography and atomic force microscopy groups and with molecular biologists and investigators engaged in other biological disciplines.
Candidates should have a Ph.D. in Physics, material science or engineering and be familiar with computer programming, instrumentation and, preferably, experience with electron microscopy and electron energy loss spectroscopy. Applications with biographical sketch, bibliography and the names of three references should be sent to: Dr. Andrew P. Somlyo, Department of Molecular Physiology and Biological Physics, University of Virginia, P.O. Box 10011, Charlottesville, VA 22906-0011, USA. The University of Virginia is an Equal Opportunity/Affirmative Action Employer.
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} Date: Mon, 9 Jun 1997 09:54:07 -0500 } To: HILDEGARD CROWLEY {hcrowley-at-du.edu} } From: Tom Phillips {tphillips-at-biosci.mbp.missouri.edu} } Subject: Re: TEM:Help!Correct PBS formula???? } Cc: } Bcc: } X-Attachments: } "Saline" is a physiological term for 0.9% NaCl solution which is 0.154 M. This is osmotically correct for human serum. If you add a large amount of phosphate buffer (e.g., 0.1M), one would have to lower the NaCl concentration in an equivalent manner if you want to maintain osmolarity. The Gibco catalog gives two formulations for PBS on page 2.9 of their 1997 catalog. one lists 9.0 g NaCl, 0.795 g NaH2POH-7H2O and 0.144 g KH2PO4 (all per liter). this is about 3 mM Na2PO4 and 1 mM KH2PO4. the other formulation is 0.21 g KH2PO4 and 0.726 g NaH2PO4-7H20. either one causes a slight hyperosmolarity in respect to straight "saline". In osmicated tissue, osmolarity might not be important but I suspect in aldehyde fixed tissue it can be. In TEM, one has to worry about PO4 based buffers causing precipitation of Ca salts. In procedures like DAB, one has to ensure the amount of buffer capacity is not exceeded. } } } All over our building we have an argument raging as to the correct } } formulation for PBS. Mainly the argument consists of opinions that the } } phosphate concentration should be 0.1M, and other opinions that the } } concentration ought to be O.OlM. (We all agree that NaCl concentration } } is 0.15M). There are also conflicting statements in the literature as to } } the composition of PBS. } } What we need is a thorough explanation of why one or the other } } concentration of phosphate salts is correct for mammalian tissue for TEM } } in procedures such as the DAB process. } } I would so much appreciate help in logically settling this argument around } } here. } } So long, } } Hildy } } } } Hildy Crowley } } University of Denver } } 2101 E. Wesley Ave } } Denver, CO 80208 }
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)
Do any of you folks have any experience using the Polaroid MicroCam with a Nikon Labophot-2 or equivalent microscope with Color-Free objectives? The MicroCam replaces the eyepiece of the microscope; it is unclear to me whether or not the MicroCam includes compensation for lateral chromatic aberation (also known as chromatic difference of magnification). The Nikon (as well as other recent microscope) objectives are corrected to provide a color free intermediate image. If the MicroCam includes compensation, it seems to me that this might negatively affect the resulting photomicrographs.
I have been unable to obtain any information from Polaroid telephone technical support on this question.
Does anyone have any experience with using a Pixera digital camera for routine light microscopy work? If so could you share your experiences/feelings/comments either to the list or off-line?
What I'm looking for:
(1) a means of directly capturing digital images from routine light microscopy (i.e. bright field, phase etc.) but not barely even visable fluorescence (i.e. no FISH!).
(2) Speed is not a factor, as we aren't interested in highend "live" video, just static images, and scan times of up 1 minute would not be a problem.
(3) Resolution is a factor: i.e. really would like better than tv resolution (i.e. 640x480), beter than 1024x1024 would be perferable.
(4) Must be color.
(5) PC systems only (i.e. Pentium Pro/PCI/Windows NT), but PCI, or SCSI is o.k..
(6) And of course $$ is always a factor (yes, I'd love a microlumina, but I only have ~$1,500 to spend on camera).
The pixera seemed like a really good compromise on resolution and price (ah, but theory and reality are two different things).
Thank you in advanced.
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."
I would like to announce the following open position; which is sepearte from and in addition to the one I posted for a senior analyst/microscopist on 4/23/97. Please spread the word-Thanks
A position is currently open for an analyst at the North Carolina State University Analytical Instrumentation Facility (AIF).
Duties and responsibilities include day to day operation and maintenance of a Cameca IMS-6F SIMS instrument, stylus profilometers, and sample preparation devices such as plasma metal coaters, vacuum ovens, etc; assistance with scheduling of access to and oversight of the above instrumentation, assistance with the teaching of surface analytical laboratory classes and assistance with other graduate level engineering classes. Requirements include a BS or higher degree in analytical chemistry or a materials related discipline (non biological) along with some hands on analytical experience, equivalent to the MS or PhD level. Experience in a multi-user SIMS facility highly desireable. Experience in vacuum equipment and/or electronics maintenance; experience with PC and Unix; and experience with analysis of semiconductor or other non biological materials a plus.
Please send resume, salary history and names of references to: Prof. Phil Russell,Director Analytical Instrumentation Facility; North Carolina State University; Box 7531, Room 118A EGRC; 1020 Main Campus Drive; Raleigh, NC 27695-7531
Email application is encouraged to prussell-at-ncsu.edu. Fax 919 515 6965
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
In my experience the Polaroid MicroCam has several drawbacks that render it useless, at least for my applications, low and meduim mag of stained mammalian histological specimens.
1. Resolution is poor on the film no matter how carefully one focuses througth the camera.
2. Color reproduction of typical biological dyes (hematoxylin, eosin, analine blue, orange G, PAS, etc) is very poor.
So much for your concerns about chromatic abberation!
3. The camera only takes (expensive) Polaroid print film so you won't have a negative to print or a slide to project.
4. Exposue varies with the specimen so many test shots are necessary.
The results I saw at two different demos were so poor that if I had a free MicroCam and a free lifetime supply of Polaroid film, I still would not use it! I was amazed that Polaroid would try to sell such a piece of junk!
Geoff -- *************************************************************** Geoff McAuliffe, Ph.D. Neuroscience and Cell Biology Robert Wood Johnson Medical School 675 Hoes Lane Piscataway, NJ 08854 voice: (732)-235-4583; fax -4029 e-mail: mcauliff-at-umdnj.edu ***************************************************************
A very useful paper to help choose the best buffer concentration and how much of common additives is: MD Maser et al (1967), Relationships among pH, osmolality and concentration of fixative solutions, Stain Technology 42:175-182. It also describes why it is important to check the osmolality and has a useful list of further reading. Diana.
Diana van Driel Dept Ophthalmology Sydney University C09 AUSTRALIA 2006
- Antique or scientific instrument dealers as well as Sotheby's or Christie's or any other local auction house should be able to value such a rarity.
- The microscope journal "MIKROKOSMOS" Stuttgart (Gustav Fisher Verlag) also publishes "microscopes for sale".
- A brass microscope in excellent condition of LEITZ or ZEISS from early this century could cost about 1500 to 2000 DEM, or $900 to $1500 US, depending on the condition and sorts of accessoires.
- Microscopes from individual manufacturers are in general lower, but estimated on a more subjective basis between nothing, 250$ and very much more than 2000 DEM. If a microscope is not from a famous manufacturer, optics are generally estimated as comparably poor, which in the case of this microscope is more or less the case: it is not possible to focus more than about 60% of the field of view at the time.
Some were concerned why I would sell such a device which was manufactured by an ancestor. I did not want to upset anyone. So if you have time and if you are interested:
My great-grandfather was a very passionate precision mechanic with an unusual high professional pride. That must be why he passed his spare time building tons (to give you an idea, the approximate cost of only having the completely unusable part of his tools put away two weeks ago were about 1000 DM) of devices like a precision scale, microscopes, a telescope, carved wooden statues, building furniture, as well as countless oil paintings etc. etc. Also, he was known to having sold *his* ancestors things to be more mobile, as he was not the first one in the family to build things together.
Rather, we seem to be a whole tribe of such persons dedicated to the combination of anything with manual work. My grandfather (mechanic, passionate builder of exquisite wooden furniture still in use throughout the family, who carved some of the family into statues), my mother (certified master of graphoanalysis I.G.A.S., passionate manual worker with beautiful oil and water color paintings to entirely fill our house, wood, cloth etc.), and her sister (a physicist who worked in the field of semiconductor crystal epitaxy and earned an IBM research prize for that, passionate piano player), and her brother (electrician, passionate oil painter), and me (on my way to forensic pathology, passionate oil painter, wood carver, playing guitar, at high-school I was doing technical and comic drawings as well as giving guitar lessons aside to earn some money; in my first clinical year, I stitched together among others a complicated chainsaw skin lesion of about 2 x 4 cm using about 50 several stitches, which healed without complications) and my sister (physical therapist, working more with cloth and clay: you just should see her flat!), as well as our cousins (one engineer and draughtsman and passionate violinist who just played in a cathedral at Einsiedeln here in Switzerland, one also physicist doing semiconductor technology, two others medical doctors) all must have some of those genes: we are all passionate about manually working in our most interesting fields, we all virtually have the inability to do other in our spare time than to carve, paint, play instruments etc. and each amass our specific load of manufactured products, each in his particular area. For the family of my father: My father is working at the Robotics department here, his brother is an architect and passionate painter as well. Other relatives: my other great-uncle was a Saddler/upholsterer who built his small store into a Prime furniture house, and the one great-uncle who died at the age of 102 incredibly enjoyed playing piano until he was at least a century old, his daughter was a professional seamstress with her own business. But we all have also the strong wish to give at least some of our ancestors devices away for more mobility.
Also I just realize that we are continuing this tradition: My girl friend (physical therapist with back-breakingly stuffed schedule despite rising health cost, passionate wood carver as well who finished her first cat statue last summer) and my sister's husband (surveyor and passionate mechanic) also love their heavily manually oriented work, as well as their relatives do (examples: one is a mechanic producing technical solutions for his firm, then, one is a medical doctor in the field of orthopedics).
Because I have been asked by some distant relative, who would very much love to buy and exhibit one of his microscopes in her living room, we are probably going to offer this item as a gift or for a reasonably low price. Surely we would not dishonor our ancestor in any way !
As our family history goes back to about 1640, I am very grateful they were such great people about their work and hobbies and everything, as am I and all around here, but I am actually quite glad they did not keep *all* those things they all happily put together primarily out of the satisfaction just to do it, and I am especially glad they are doing Quality Control over there at LEITZ's and ZEISS'.
And I especially acknowledge your help and understanding, Wolf Schweitzer
I am an Information Research Specialist with Motorola SSTG in Scottsdale, Arizona. I am having difficulty locating a certain document and thought this list might be of assistance in my search. Any help or suggestion is greatly appreciated.
The document I am looking for is :
"Auger, EMPA, XPS, and EBSP Analysis of Discolored Au/Ti/Cu Multilayer Thin Film I/O Pads", R. E. Davis, J. L. Hurd, J. Gates Jinkins, and R. Flitsch, Microbeam Analytical Society, Aug. 1993, Los Angeles, CA
If you know of this document and where I may obtain it, please contact me via email directly at p28167-at-email.mot.com.
Thank you in advance for your assistance and best regards,
Donald Guy
Information Research Specialist SSTG Learning Center/Library Motorola Space and Systems Technology Group 8220 E. Roosevelt St., MD R1206 Scottsdale, Arizona 85252
We have been using a Web-based sign-up system for our confocal for about a year and like it very much. The system can be seen via the "Confocal Schedule" link on the following Web-page:
http://ww2.med.jhu.edu/confocal/
Only registered (i.e. properly trained) users can sign up, using their password, up to one month in advance. The system is such that any person's time can be cancelled by any other user in their own lab -- very handy in the case of illness, experiments going longer than expected, or the boss informing you that you really should be working on project X rather than Y. The facility does charge a "no-show" fee for folks who reserve time and don't use it. This is a putative measure that can always be avoided (even for last minute changes in plans) by informing the person in charge of billing and the person signed up after you.
Greg Martin Dept. of Cell Biology and Anatomy Johns Hopkins School of Medicine
On Wed, 4 Jun 1997, Tamara Howard wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Does anyone have on-line sign-up for equipment time? I remember this was } discussed here ages ago, but I haven't heard about it recently. We are } thinking about going to such a system, and would like to hear } pros/cons/horror stories/suggestions. } } Thanks! } } Tamara } CSHL (NY) } howard-at-cshl.org } } }
Hello all, What is the best way to process cells for TEM which are grown on glass coverslips affixed to plastic petri dishes. I will need to remove the glass from the plastic and cut ultra thin sections. I have previously tried to do this and could not remove the glass. I gave up on glass and started using aclar. I now must return to using glass. Any suggestions?
Sometimes if all else fails, I take a wood stick, dip it in to chloroform and wave it over the sections in the boat like a magic wand. If its not an infiltration problem, this can flatten the sections.
Bob U of Washington Seattle
On Mon, 9 Jun 1997, William Meek, Ph.D. wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } I am having problems with wrinkles in Epon thin sections (gold } interference colors)and would like some advice or information about } getting rid of them. The specimen is ameba and has been grown on the } surface of an Epon bullet coated with a collagen matrix. The ameba are } then fixed, dehydrated and embedded within the Epon bullet. Early } sections (the ones first off) are of interest since the ameba tend to } attach to substrate. } } Bill }
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We will be receiving an FTIR with a microscope accessory soon. I would appreciate comments about this technique from users whose application is drug identification or fragment ID of particulates sometimes found on food. Does it truly have the power to zero in on the suspect substance without sample extraction/cleanup? Formerly we used diffuse reflectance or made pellets for FTIR spectra of our old spec without a microscope. Thanks chemist SLI
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Sally: Our EM facility has been using glass coverslips in several tissue culture experiments. After flat embedding with epoxy/alradite resin, we remove the glass, using cold HF[hydroflouric acid] (in a Fume Hood!) for -at-4-8 minutes. We get 100% success rate, using many different types of glass coverslips. However, the glass cover slips are not attached to plastic petri dishes. Do the coverslips need to be attached to petri dishes? The glass surface has to be exposed to the HF fluid. You can still use this technigue if your coverslips are atteched. you will have to add an extra step to remove the glass from the plastic, via sandpaper. Please contact me, if you would like a detailed step by step procedure for this technique. It is very straightforward.
The references we started with: Moore, M.J., 1975, Removal of Glass Coverslips from Cultures Flat Embedded in Epoxy Resins Using HF, . Microscopy, v. 104, p. 205.
Rieder, Conly and Bowser, Samuel, 1987, "Correlative LM and EM on the Same Epoxy Section", in Correlative Microscopy in Biology , Chapter 11, Academic Press, 1987.
-Louisa howard Dartmouth College EM Facility Hanover, NH. 03755 (603) 646-3492 Louisa.Howard-at- Dartmouth.edu
We are using some classical light microscopy staining techniques to screen our plant samples before we carry out EM on them. Many of these techniques include carrying the samples through xylene baths. We have been encouraged to find a safer alternative, if possible. I remember that years ago a xylene replacement was introduced into the marketplace to be used during paraffin embedding of samples, but I heard that this was not a good replacement, and it was discontinued. I don't know what happened after that or what is used these days.
Any help on this matter would be greatly appreciated. Thanks.
Paula.
Paula Allan-Wojtas Food Microstructure Specialist Agriculture and Agri-Food Canada Atlantic Food and Horticulture Research Centre Kentville, Nova Scotia Canada B4R 1A1
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Hello all, } What is the best way to process cells for TEM which are grown on glass } coverslips affixed to plastic petri dishes. I will need to remove the } glass from the plastic and cut ultra thin sections. I have previously } tried to do this and could not remove the glass. I gave up on glass and } started using aclar. I now must return to using glass. Any suggestions? } } Sally } }
Group A reader of mine, not on this listserver, is "interested in obtaining electron micrographs of a number of microbes: T. Pallidium, S. pneumoniae, H. pylori, V. Cholerae, m. tuberculosis, and others." Should you be able to help, please contact Bruce Cameron directly at: bcameron-at-tigr.org Regards to all, Don Grimes, Microscopy Today
The newest version of the Image Processing Tool Kit is now completed. The CDs are being sent free of charge to upgrade all registered users of the kit. Version 2.1 includes about 80 Photoshop compatible plug ins with image processing and measurement functions. Many of the plugins offer speed improvements of 2x to more than 10x over version 1, and are fully compatible with the Layers structure used in Photoshop 4. The Mac and PC versions (both on the CD) are the same. The CD includes a much expanded tutorial (175 pages) and more than 100 test images. It also has versions of the tutorial that specifically discuss the use of the tool kit with NIH-Image (Mac) and ImageTool (Win). Full info on the package is available from http://members.aol.com/ImagProcTK/ A review of the original CD is available on-line from http://www.macscitech.org/stj/stj1997_apr/stj1997_apr.html#one
Jim, I tried immersing my plastic in liquid nitrogen and was unable to remove the glass. Perhaps you did something special during the whole process of embedding, or perhaps I am not using the best plastic. I use embed 812 kit. Also, I have heard that if you use serum in the culture medium, then the glass will not come off the surface of the plastic. Does this make sence?
Thanks for the info on Histoclear. I've been using a supply that was=20 here before me and I wasn't sure if it was available anymore or where to=20 get it. It's interesting to read that it's not for use with paramount- I= =20 use it all the time, even use the Histoclear to thin the paramount when=20 it starts to get too thick. I haven't noticed a big problem, although it= =20 takes the coverslip alot longer to dry down. In one of my applications,=20 I rinse celloidin sections in Histoclear prior to pressing them flat on a= =20 slide and trimming the edges of the section with a razor blade. I like=20 Histoclear better for this because is does take a while to evaporate and=20 gives me more time to work with the sections before they dry out. (They=20 need to stay moist throughout the mounting process.) =20
One more note. Althought Histoclear is supposed to be nontoxic, it gives= =20 me a headache, so I always work in a hood.
Karen
On Tue, 10 Jun 1997, Philip Oshel wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } =20 } } We are using some classical light microscopy staining techniques to } } screen our plant samples before we carry out EM on them. Many of these } } techniques include carrying the samples through xylene baths. We have } } been encouraged to find a safer alternative, if possible. I remember tha= t } } years ago a xylene replacement was introduced into the marketplace to } } be used during paraffin embedding of samples, but I heard that this was } } not a good replacement, and it was discontinued. I don't know what } } happened after that or what is used these days. } } } } Any help on this matter would be greatly appreciated. Thanks. } } Paula Allan-Wojtas } =20 } Paula, } =20 } There is a fine replacement available (unless it's been taken off the } market in the last year): HistoClear, marketed by National Diagnostics. } This is also sold by Fisher, or if they don't have HistoClear, they have } their own brand of the same thing. } =20 } It's "essential oils of citrus", and smells like oranges. Non-toxic, work= s } very well for paraffin sectioning, or any other procedure that uses } xylenes. But (there's always a but), you can't use Permount or the like t= o } mount the coverslips. N. D. has a replacement product HistoMount (as does } Fisher, I believe). This works very nicely, But ... , you can't warm the } slides to set the mountant as you do with Permount, etc. Otherwise the } HistoMount shrinks like mad. Shrinks some anyway, but this in only a majo= r } annoyance if you're doing very thick sections. The usual 10-15=B5m ones a= re } no trouble. } =20 } Note: companies are mentioned because I think National Diagnostics } developed (or first commercialized) this stuff, and because I know they a= nd } Fisher sell it. Other companies probably do as well. (N.D. also came out } with a replacement for foraldehyde as a fixative, using a sugar aldehyde. } This didn't seem to work so well, and *maybe* has been withdrawn, but I } don't know.) } =20 } Phil } =20 } } Sic Hoc Legere Scis Nimium Eruditionis Habes { } 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 ****************** } =20 } =20 } =20 } =20
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Subject: Time:7:32 AM OFFICE MEMO Post-doc position at LLNL Date:6/11/97
Our Center for Accelerator Mass Spectrometry (CAMS) is currently seeking a Postdoctoral Research Staff Member to work as part of an interdisciplinary team of physicists, chemists, and biologists. In this research position, you will work as part of a team to develop quantitative elemental analysis of biological tissues. These techniques will be used in studies of elemental kinetics, structural biology, toxicology or pharmacology by staff at LLNL and collaborators in the University of California system and other institutions. Duties include development/modification of techniques for preparation of biological tissues for quantitative elemental microanalysis, operation of the LLNL microprobes to analyze prepared biological samples, X#030#ray data reduction and analysis, planning/designing and executing independent and collaborative research projects and publication of results in peer#030#reviewed literature. Requires a recent Ph.D. in chemistry, biochemistry, toxicology, pharmacology or related field. Experience in trace element analysis and analytical microscopy is desired. LLNL offers a challenging work environment and a competitive salary/benefits package. Qualified individuals are invited to send their resume to: Lawrence Livermore National Laboratory, Attn.: Mary Anne Holman, L#030#725, P.O. Box 5510, R#030#4249, Dept. AJSC527MH, Livermore, CA 94551#030#5510. LLNL is an Equal Opportunity Employer. Lawrence Livermore National Laboratory
I have been using micro-FTIR for years to identify particle and fiber contaminants in pharmaceutical products. Your statement about being able to identify particulates without extraction is partially true; it depends alot on the matrix in which the contaminant is found. I have found it useful to at least "rinse" the contaminant in a solvent (in these applications the solvent is usually water) first. Otherwise, you may have trouble with minor extraneous absorptions which hamper the positive identification of your contaminant, especially with automated search routines.
I have found that a low power (10-60x) stereomicroscope with dark field lighting is imperative in the pre-analyis prep of such samples. A clear glass spot plate (or microscope slide with wells) can be used for extraction/cleanup of your particle. You can view it under the 'scope while the extraction occurs. You also should have a good supply of micro-probing tools (available from most SEM supply houses) if you don't already have them. If you "tease" the particle around in the solvent, and pull it away from the solvent as it evaporates under the heat of the 'scope you can usually clean up the particle enough to obtain a good spectra.
Particle size is another consideration. Typically, you can obtain a useful transmission spectrum from particles greater than about 20 microns. Reflectance spectra may require larger sizes. Fibers generally work quite well in transmission. In fact, I once built a library of spectra for the types of fibers found in the cleanroom garments used in the compounding area for a pharmaceutical product. I subsequently used that library to identify/troubleshoot particulate problems for the customer.
I hope this info is useful to you. Good luck in your identification work.
Regards,
Bob ***************************** Bob Citron Chiron Vision Corp. Claremont, CA 91711 USA (909)399-1311 Bob_Citron-at-cc.chiron.com *****************************
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We will be receiving an FTIR with a microscope accessory soon. I would appreciate comments about this technique from users whose application is drug identification or fragment ID of particulates sometimes found on food. Does it truly have the power to zero in on the suspect substance without sample extraction/cleanup? Formerly we used diffuse reflectance or made pellets for FTIR spectra of our old spec without a microscope. Thanks chemist SLI
X-Authentication-Warning: duchesse.zdv.Uni-Mainz.DE: windoff owned process doing -bs
Hello, I am working with cells growing on round cover slides. We are looking for an apporbiate holder to handle them in larger amounts during immunohistochemistry. Something like a smaller version of the staining jars which are used for normal microscopic slides. Can someone help me?
I use a very nice ceramic coverslip holder purchased from Thomas Scientific. You can also purchase glass jars of the appropriate size from the same catalog. Note that this item may be available from other vendors as well. This holder carries a dozen coverslips and has a metal handle that you can remove. It's actually a miniature version of the regular slide holders. I used it with square coverslips but regular sized round covers should hold without problem. Sorry, I don't have the current reference at hand. Good luck, 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. ****************************************************
} Hello all, } What is the best way to process cells for TEM which are grown on glass } coverslips affixed to plastic petri dishes. I will need to remove the } glass from the plastic and cut ultra thin sections. I have previously } tried to do this and could not remove the glass. I gave up on glass and } started using aclar. I now must return to using glass. Any suggestions? } } Sally
Have you tried using Thermanox coverslips? Not cheap, but they are good for culturing and good for embedding and they peel away from resin with the application of a little heat from a hot-plate. Thermanox can also be sectioned along with the resin if you like, but I have had problems with this because the Thermanox and resin do not stretch to the same degree, so you end up with sections wrinkled on one edge.
We have "popped" cells grown in both serum and serum-free medi on glass coverslips using liquid nitrogen 100's of times. serum makes no difference. The coverslips are fixed either in the 12 well plastic trays or transfered to 20 ml glass scint vials. by the time we get to the dehydration steps, we always switch to the glass vials. after 100% ethanol, we infiltrate with embed 812 (1:1 with ethanol overnight) then 100% resin for 3 hrs then place the coverslip cell side up on a glass slide with the thin layer of resin that comes with it when we transfer to the slide (you don't want it too thin or too thick but it if you are in doubt, simply add 1 drop to the coverslip after transfering and let it flow naturally. you shouldn't get a final plastic spread of more than 1.5 - 2 times the diameter of the coverslip or you are using too much. when you take the slides out of the oven, let them cool for 15 min and touch with a razor blade. if you get a gooey strand when you pull the blade away, you haven't polymerized it enough. if it is not too gooey, cross-hatch the surface of the plastic using the razor blade to score deeply into the plastic (to the level of the glass). SLOWLY immerse the slide into Liquid N2 and the squares should pop right off. look at the surface of the coverslip and bottom of the square to ensure no glass is going with the square. if so, you have over polymerized. for some projects, we simply put the square in the flat microtome holder and cut it but ususally we re-embed the square in a standard mold. if you aren't re-embedding, you should heat the squares in a rubber mold overnight. once you have the timing of how long to polymerize the coverslips, it is trivial. we havent had a failure in years of doing this a lot of times. good luck.
Tom Phillips
} Jim, } I tried immersing my plastic in liquid nitrogen and was unable to remove } the glass. Perhaps you did something special during the whole process of } embedding, or perhaps I am not using the best plastic. I use embed 812 } kit. Also, I have heard that if you use serum in the culture medium, then } the glass will not come off the surface of the plastic. Does this make } sence? } } Sally
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)
This fall and winter I need to fix and embed dormant blueberry buds; it seems likely that I will run into problems trying to infiltrate the somewhat woody exterior with fixatives and the embedding medium. We're going to be immunolabeling so we'll be using LR White but I'm wondering if anyone with experience with woody tissue can recommend buffers and fixatives ?
We generally use 0.2% glutaraldehyde, 3% paraformaldehyde in a 0.05M buffer and this works fine for non-woody tissues so I thought I'd start there but I would greatly appreciate any advice!
} there that can do this faster. Does anyone know of such a software? I } would appreciate comments.
GLOBAL LAB (Data Translation)
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I've been asked to help some colleagues monitor the production of hydrogen peroxide in potato leaves undergoing bacterial attack. I have a paper citing a technique using CeCl3 - cut samples are incubated in the cerium prior to fixation (glutaraldehyde, paraformaldehyde, sodium cacodylate buffer , post-fixed in osmium), dehydration (ETOH and propylene oxide) and embedding (Eponaraldite). I've never done this before and am wondering: can anyone give me some helpful tips, hints, advice etc?
Hi, folks. I've inherited an Orthomat camera (old-style, not the more recent Wild-Leitz type) without the film cassette. If you have one or more that you'd be willing to part with for $$ or barter, please contact me at smithj-at-winthrop.edu. TIA Julian
Julian P.S. Smith III Biology Winthrop University Rock Hill, SC 29733 803-323-2111 x227 (vox) 803-323-2246 (fax)
I have come across the use of 'Histoclear' - sorry that's all I know about it. But in recent years we have used an even safer alternative called 'Citroclear' (I believe its an extract of citrus fruits) - in the UK this is supplied by: HD Supplies 44 Rabans Close Rabans Lane Industrial Estate Aylesbury Bucks HP19 3RS UK Phone Aylesbury (01296) 431920 - I will leave you to work out the international dialling codes Fax Aylesbury (01296) 392121
The cost in our 1995 price list was: catalogue number - CC500 Citroclear 5litres - 16.75 UK pounds 25litres - 74 UK pounds the prices are ex VAT(purchase tax) ex delivery and probably out of date.
It appears to do much the same job as xylene but is ( at this present time) considered to be a lot less toxic. It's hazard data sheet lists it's known hazards as an irritant with the possibility of dermatitis after long exposure. Other characteristics are a strong fruity smell which some people like and some don't and a tendency to turn yellow and throw out oily deposits with prolonged storage in light. Oh the suppliers say it is bio-degradable as well and can be carefully disposed of down the drains if your regulations allow.
I don't know if you can source this in Canada but good luck in your search. By the way I have never used xylene outside of a fumehood in years.
Malcolm Haswell University of Sunderland UK
Disclaimer - I have no connection with the company other than as a satisfied user.
----------
Separating glass from plastic is easy in my experience after thermal shock. Immerse the specimen in liquid nitrogen for a few seconds. 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
} From: Sally Shrom {sally-at-retina.anatomy.upenn.edu} } To: Microscopy-at-sparc5.microscopy.com } Subject: TEM cell culture } Date: Tuesday, 10 June 1997 23:13
} } Hello all, } What is the best way to process cells for TEM which are grown on glass } coverslips affixed to plastic petri dishes. I will need to remove the } glass from the plastic and cut ultra thin sections. I have previously } tried to do this and could not remove the glass. I gave up on glass and } started using aclar. I now must return to using glass. Any suggestions? } } Sally
If the usual stretching with solvent or heat (which is required to compensate the compression artifact) does not do the job, try using a loop to pick up the section. Then deposit these onto a grid which is sitting on filter paper. This solved a wrinkle problem I had with thick cell walls. 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
} On Mon, 9 Jun 1997, William Meek, Ph.D. wrote: } I am having problems with wrinkles in Epon thin sections (gold } interference colors)and would like some advice or information about } getting rid of them. The specimen is ameba and has been grown on the } surface of an Epon bullet coated with a collagen matrix. The ameba are } } then fixed, dehydrated and embedded within the Epon bullet. Early } sections (the ones first off) are of interest since the ameba tend to } attach to substrate. } } Bill
} We are using some classical light microscopy staining techniques to } screen our plant samples before we carry out EM on them. Many of these } techniques include carrying the samples through xylene baths. We have } been encouraged to find a safer alternative, if possible. I remember that } years ago a xylene replacement was introduced into the marketplace to } be used during paraffin embedding of samples, but I heard that this was } not a good replacement, and it was discontinued. I don't know what } happened after that or what is used these days. } } Any help on this matter would be greatly appreciated. Thanks. } Paula Allan-Wojtas
Paula,
There is a fine replacement available (unless it's been taken off the market in the last year): HistoClear, marketed by National Diagnostics. This is also sold by Fisher, or if they don't have HistoClear, they have their own brand of the same thing.
It's "essential oils of citrus", and smells like oranges. Non-toxic, works very well for paraffin sectioning, or any other procedure that uses xylenes. But (there's always a but), you can't use Permount or the like to mount the coverslips. N. D. has a replacement product HistoMount (as does =46isher, I believe). This works very nicely, But ... , you can't warm the slides to set the mountant as you do with Permount, etc. Otherwise the HistoMount shrinks like mad. Shrinks some anyway, but this in only a major annoyance if you're doing very thick sections. The usual 10-15=B5m ones are no trouble.
Note: companies are mentioned because I think National Diagnostics developed (or first commercialized) this stuff, and because I know they and =46isher sell it. Other companies probably do as well. (N.D. also came out with a replacement for foraldehyde as a fixative, using a sugar aldehyde. This didn't seem to work so well, and *maybe* has been withdrawn, but I don't know.)
Phil
} Sic Hoc Legere Scis Nimium Eruditionis Habes { 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 ******************
} } I am having problems with wrinkles in Epon thin sections (gold } } interference colors)and would like some advice or information about } } getting rid of them. The specimen is ameba and has been grown on the } } surface of an Epon bullet coated with a collagen matrix. The ameba are } } then fixed, dehydrated and embedded within the Epon bullet. Early } } sections (the ones first off) are of interest since the ameba tend to } } attach to substrate.
Usually the first sections to come off of organisms growing on surfaces tend to be somewhat problematic probably due to the the irregularities of the surface and the modifications of the cell that facilitate adherence to the surface. Such sections tend to wrinkle and may split apart in some cases. We find that picking up the sections onto a naked grid (flamed nickel grid, for example) and passing the sections (still floating on a droplet of water on the grid) between a heated electrical loop will tremendously relax the sections - more so than chloroform. If this does not work, then consider varying the clearance angle of the knife as well as the sectioning speed, increasing the hardness of the plastic, or use a good, smaller angled (or diamond) knife. Good luck.
#################################################################### 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 ####################################################################
Are your sections dried down onto a formvar or other support film? Wrinkles can also occur when sections "loosen" during the staining process and then subsequently dry down again (unevenly). This can be remedied by firmly attaching the sections to the support film by heating the grids at 60 C for 15-30 min. This is best done using a heating block. Good luck.
Tim Bourett DuPont Experimental Station Wilmington, DE USA
I am seeking to purchase used, the epi-fluoresence accessories for a Leitz Orthoplan that is over 20 years old.
Please reply privately.
Thanks, Greg Erdos ******************************************************* 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
We have used FTIR microscopy routinely in our labs for a few years and are enthusiasticly for it. Two of the most important advantages are the simple sample preparation and the ability to take spectra of small particles. We often wish to find polymorphs (which may be a small percentage of the whole) of investigational drug substances and FTIR allows us to look at individual particles. Its great for aiding in the identification of small bits of polymer contaminants which can be tough by polarized light microscopy.
One of the limitations is a familiar one to all of microscopy - sampling. How can one know whether the selected particle is truly representative of the whole? We also think it is necessary to crush the particle to reduce IR polarization effects. The minimum size given by some of the manufacturers also seems a bit small in normal applications. We like particles in the 30 to 70 um region.
Our routine sample preparation technique involves mounting the particle or particles on a KBr window under a stereomicroscope. We then gently crush the particle with a stainless steel roller. We then either mark the location or keep in mind the surroundings. One problem we've had is finding the particle of interest using the optics of the IR scope. We are then ready to test.
I have a big problem, osmium precipitates!! does anyone can help me? Thanks in advance Nuria Cortadellas Department of Electron Microscopy University of Barcelona
I'm sorry I can't address the question about the corrections of the Polariod Micro camera but I would like to respond to the torching it received. I agree that the results are far from critical photomicrography, but it does have a niche in my lab. I have a stereomicroscope without a camera and there are times that I require simple, quick photo documentation. For instance, I just worked on some tablets with unsightly spots and I wanted to document their location and color before I analyzed the spots by EDS. The Polaroid Micro worked quite nicely. The thing I like the most is that it is an SLR so I get to see exactly what I'm photographing and can judge focus easily. Its surely not a Cadillac and probably not a Chevy but it may be a Moped.
} Hello, } I am working with cells growing on round cover slides. We are looking for } an apporbiate holder to handle them in larger amounts during } immunohistochemistry. Something like a smaller version of the staining } jars which are used for normal microscopic slides. Can someone help me? } } Thanks } Reinhard } } . . . . . . . . . . . . . . . . . . . } Dr. Reinhard Windoffer Fon: (00)49 (0)6131/39 3720 } Universitaet Mainz Fax: (00)49 (0)6131/39 4615 } Anatomisches Institut e-mail: windoff-at-mail.uni-mainz.de } Becherweg 13 } D-55099 Mainz } Germany } . . . . . . . . . . . . . . . . . . . Reinhard, I have made round coverslip holders from fairly thick walled flexible polyethylene and silicone tubing by cutting it in half lengthwise and then cutting slits perpendicular to the length. When the tubing is flexed in the correct direction the slits are open for inserting the cover slips. When the tubing is held straight (in a pyrex tube or syringe barrel) the slits pinch closed and hold the cover slips. Use a tubing diameter near that of the cover slips. A cautionary note, if you plan to critical point dry samples held this way, be advised to test the tubing material in the CPD unit first. Some tubing types will turn into foam during CPD.
good luck
cheers
ed
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
} } Hello, } I am working with cells growing on round cover slides. We are looking for } an apporbiate holder to handle them in larger amounts during } immunohistochemistry. Something like a smaller version of the staining } jars which are used for normal microscopic slides. Can someone help me? } } Thanks } Reinhard } } . . . . . . . . . . . . . . . . . . . } Dr. Reinhard Windoffer Fon: (00)49 (0)6131/39 3720 } Universitaet Mainz Fax: (00)49 (0)6131/39 4615 } Anatomisches Institut e-mail: windoff-at-mail.uni-mainz.de } Becherweg 13 } D-55099 Mainz } Germany } . . . . . . . . . . . . . . . . . . . Reinhard, In my previous response I forgot to mention that a company here in the US, Tousimis Research Corp. also sold, at one time, a metal holder designed for CPD of stacks of cover slips. I have used it with good results for CPD. They come in two sizes. 13mm and 25mm? Ph: 800-638-9558 or 301-881-2450. (No affiliation, only a satisfied customer)
cheers ed
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
After reading the Glutaraldehyde/formaldehyde safety discussion, I've decided to add my note of warning. After working with GAL and formaldehyde and other chemicals of the TEM trade for 30 years with only slight reactions to them, I was exposed to a major formaldehyde spill. Not only did I suffer major respiratory tract irritation, but my lung collaped and I soon developed hypersensitivity to a wide range of volatile organic chemicals, including formaldehyde, GAL, liquid epoxy resins, solvents, 2-mercaptoethanol, pesticides (including herbicides), auto exhaust, etc. When exposed to these I get headaches and "fuzzy" in the head almost instantly, and a delayed hypersensitivity reaction in my pleural cavity about 30' to 2 hr later that is extremely painful. This lead to further pneumothoraces until I had lung surgury. Our hypothesis is that I have developed an immune response to formaldehyde-altered proteins of my lung or pleural cavity, as well as the limbic system (neural) response often seen in others with chemical hypersensitivity.
Needless to say, I recommend keeping your exposures to a minimum. All use of aldehyde fixatives should be in a well-functioning fume hood. Anatomy classes should have individual fume hoods for each dissection station and/or switch to non-aldehyde fixed cats for example (we did both). Incidently, my unofficial survey of former occupations of persons with MCS indicates that histology technicians have the highest incidence and some formal surveys show similar results.
It's definitely less fun trying to function (and very difficult to do research) in today's chemically-dependent society when you are hypersensitive to nearly every volatile organic chemical around. -BE CAREFUL WITH THE GAL AND FORMALDEHYDE!-
-Dennis
Dr. M. Dennis Goode Phone (301) 405-6917 Department of Zoology Fax (301) 314-9358 University of Maryland e-mail goode-at-zool.umd.edu College Park MD 20742 ************************************************************* "If the Lord Almighty had consulted me before embarking upon the creation, I should have recommended something simpler." -Alphonso X of Castile, 15th Century
I am trying to obtain the contour of wheat to build the three dimension model of wheat. The contour image of wheat is not good. It is not continuous. If someone has good idea or method, please let me know. I sliced the wheat using ultramicrotome with glass knife. Then I obtained the image of wheat slice. I tried to obtain the contour image using Imaging software. Thanks.
Question #1: I have a Haskris RO75 water chiller and I am in need of another; I have found ALL of the components to make it a double pumper though. Has anyone turned their single pump RO75 into a double pumper? What are the pitfalls? Am I nuts or just cheap?
Question #2: A client of mine wants to see if she can see the difference between elastin and collagin at the TEM level {she is looking for the relative amounts in various samples}. Is this possible? What are the references/protocols? Am I nuts to try this?
Thanks, John Grazul Rutgers University Electron Imaging Facility
Global Lab is no longer sold by DT. You can try Visilog from Noesis Vision Inc. We have a version available for 2,995.00$ US.
At 05:43 PM 6/11/97 +0100, Rui Costa wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
---------------------------------------------------------------------------- ------------- Luc Nocente Tel: 514 345 1400 Noesis Vision Inc. Fax: 514 345 1575 e-mail: ln-at-noesisvision.com http://www.noesisvision.com 6800 Cote de Liesse, Suite 200 St-Laurent, PQ H4T 2A7,Canada ---------------------------------------------------------------------------- -------------
I recently asked for information regarding a Zeiss SEM 6. Subsequent direct conversations with the donor clarified that, in reality, a free Zeiss 9A TEM is available from here in San Diego. It was donated to the school district and they have been awaiting funds to install it. Those funds never materialized, so the district now wants to pass it along. I have no personal knowledge of the working condition of the scope--I do know it is currently sitting in a warehouse. Anyone interested in getting this donation should contact Debbie at the Grossmont School District Instructional Services Dept. at (619)579-9711. They are going to dispose of it in the very near future.
Dr. Steven Barlow EM Facility/Biology Department 5500 Campanile Drive San Diego CA 92182-4614 phone: (619)594-4523 fax: (619) 594-5676 email: sbarlow-at-sunstroke.sdsu.edu website: http://www.sci.sdsu.edu/EM_Facility
} Hello plant pro's, } } This fall and winter I need to fix and embed dormant blueberry buds; it } seems likely that I will run into problems trying to infiltrate the } somewhat woody exterior with fixatives and the embedding medium. We're } going to be immunolabeling so we'll be using LR White but I'm wondering if } anyone with experience with woody tissue can recommend buffers and } fixatives ? } } We generally use 0.2% glutaraldehyde, 3% paraformaldehyde in a 0.05M } buffer and this works fine for non-woody tissues so I thought I'd start } there but I would greatly appreciate any advice! } } Thanks,
Peggy,
I am no plant pro, however I have had some experience with doing some plant stuff for TEM. I had a couple of users looking at grass samples and also some clover root nodules, As you would probably know, very hard to fix and infiltrate adequately. We used Microwave fixation for this purpose and also used the microwave to aid in the infiltration of the resin. Initially, this was not for Immuno but the results were far superior and in a much shorter time. We did fix some grass bits for immuno using the MW, but still had a few problems with infiltration at low temps (Lowicryl K11M). I would say that using the MW would be able to help you infiltrate your 'woody tissues' with LR White too.
Rich.
----------------------------------------------------------------------- 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!" ------------------------------------------------------------------------
I will be involved with operating an ESEM in the near future and need information about any courses available concerning this technology. Any information will be greatly appreciated.
I will be involved with operating an ESEM in the near future and need information about any courses available concerning this technology. Any information will be greatly appreciated.
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Subject: Time:7:32 AM OFFICE MEMO Post-doc position at LLNL Date:6/11/97
Our Center for Accelerator Mass Spectrometry (CAMS) is currently seeking a Postdoctoral Research Staff Member to work as part of an interdisciplinary team of physicists, chemists, and biologists. In this research position, you will work as part of a team to develop quantitative elemental analysis of biological tissues. These techniques will be used in studies of elemental kinetics, structural biology, toxicology or pharmacology by staff at LLNL and collaborators in the University of California system and other institutions. Duties include development/modification of techniques for preparation of biological tissues for quantitative elemental microanalysis, operation of the LLNL microprobes to analyze prepared biological samples, X#030#ray data reduction and analysis, planning/designing and executing independent and collaborative research projects and publication of results in peer#030#reviewed literature. Requires a recent Ph.D. in chemistry, biochemistry, toxicology, pharmacology or related field. Experience in trace element analysis and analytical microscopy is desired. LLNL offers a challenging work environment and a competitive salary/benefits package. Qualified individuals are invited to send their resume to: Lawrence Livermore National Laboratory, Attn.: Mary Anne Holman, L#030#725, P.O. Box 5510, R#030#4249, Dept. AJSC527MH, Livermore, CA 94551#030#5510. LLNL is an Equal Opportunity Employer. Lawrence Livermore National Laboratory
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At 11:45 AM 6/11/97 +0200, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Osmium which has been reduced can be reoxidised with hydrogen peroxide. Add it drop by drop until the light straw colour of OsO4 is restored. I cant see why it shouldn't be as good as new afterwards. Hydrogen peroxide is also good for wshing out a bottle before making up OsO4 solution in it.
Mel Mel Dickson President, Australian Society for Electron Microscopy Director, Electron Microscope Unit, University of New South Wales. Sydney NSW 2052 Australia
We are looking to purchase a used Carbon evaporator for SEM, EPMA work. The Denton DV-502 or 502A is a desired unit. The Denton Bench Top Turbo unit would also be highly considered. Any information would be greatly appreciated.
Thank You Harry A. Ekstrom AlliedSignal Engines M/S 46-00/302-101 P.O.Box 52181 Phoenix, AZ 85072-2181 602-231-2744
It is very easy to distinguish collagen fibers from elastin in routinely fixed and stained biological samples. Any atlas of ultrastructure (e.g. Johannes A. G. Rhodin, etc.) will have typical examples of each. Basically collagen in longitudinal section displays a characteristic 640 Angstrom banding pattern while elastic fibers have an amorphous central substance surrounded by a peripheral mantle of microfibrils. In cross-section, the two components of elastin will be visible while collagen will appear to be uniform (protein). At least that's the way I remember them from my connective tissue research days.
I was wondering if anyone knows where I can get a copy of the Transmission Electron Microscopy Monograph series by Edington. I believe they are out of print but I was hoping that there may be a place that still has a supply of them. Thanks.
Your problems with wrinkels- can be caused through many ways Are you sectioning with glass or diamond knife? It is possible to eliminate wrinkels by vapours of organic solvent (xylene, chloroform, trichloroethylene, etc) to do this for example, a sheet of filter paper or a wooden applicator stick saturated with the solvent is held as close as possible above the sections on the water surface, but without touching the sections. this should expand the sections, and get rid of wrinkels. I found chloroform to be more successful.
If you cannot expand with all solvents available - than use heat. to do this suitable wires are needed, can be bought commercially. You can use a wire loop, used for picking cryo sections 2mm diameter. Heat wire with a bunsen burner (until red hot) and than wave over sections. Plastic sections have the advantage that they can be spread by heat.
If the above does not work, than check your knife height, knife angle -check to see if specimen block , knife etc is tightened. - Specimen face propely trimmed (small face) -if you are sectioning with glass knife, try tungsten coated knife or diamond knife. ( clean knife, dirty knife will give you this problem) -If wrinkels still persists, pick up sections onto grids and view. It sometimes expands under the electron beam. - Check your epon resins it might be soft. - polmerization for epon resin in our lab. we cure for 48 hours at 70 degrees centigrate. I have vast experience in sectioning all types of resins, if your problem still exists, you are most welcome to contact me and we can discuss your problems.
I do hope some of my contributions will help to sort out your problems.
Best of luck
Vijay H Bandu Centre for Electron Microscopy University of Natal Private Bag X01 Scottsville 3209 South Africa
} Question #2: A client of mine wants to see if she can see the } difference between elastin and collagin at the TEM level {she is } looking for the relative amounts in various samples}. Is this } possible? What are the references/protocols? Am I nuts to try this? } } Thanks, } John Grazul
I've seen one response to this question already, which points out quite rightly that elastin and collagen are both quite distinctive in appearance in the TEM, and so they can't be confused with one another.
However I suspect John's problem is that neither collagen nor elastin stains well with the usual U acetate/Pb citrate combination. Here are our methods (cheap!):
For elastin - Make up 0.2% orcein in acid alcohol (0.1g orcein, 50ml 70% ethanol, 0.3ml conc. HCl) and filter. Keeps well. Stain grids on drops of this stain (e.g. on a wax sheet, covered) for 30 mins at room temp., then rinse with 50% ethanol and stain with Ua and Pbc as usual.
We use this to demonstrate the elastin in artery walls, and both newly-forming elastin and "old" elastin stains well. I don't have a reference for this method, I'm afraid ("Ancient lecipe, rost in time...").
For collagen - Make up 0.01% tannin (tannic acid) in water, preferably fresh, and stain grids on drops of this for 3 mins at 60 deg.C. Rinse with water and then stain with Ua and Pbc as usual.
Collagen shows up well, but so do some other extracellular components including elastin sometimes. The reference is: Dingemans et al. (1990) Ultrastructural Pathology ,vol.14, 519-527.
N.B. - We always post-stain with Ua and Pbc because we want to see other features in the sections, and it is possible orcein and tannin won't work on their own. So if you want to stain only collagen and elastin, and nothing else, you might have to resort to immunogold methods.
Regards
Stephen Edgar
Electron Microscope Unit, Pathology Department School of Medicine University of Auckland Private Bag 92019 Auckland New Zealand
If you are using coverslips of 13mm size you can process them in 24 well tissue culture plates.. Else the 12 well plates for larger ones.
Neelima Shah.
At 11:29 AM 6/11/97 +0200, Reinhard Windoffer wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Dear All: Steve Barlov wrote about a used Zeiss 9 from San Diego. Although I do not know this particular EM, I am quite familiar with this type. Here are some details: It is some 25 years old, has a max mag of about 60k and 60kV accelerating voltage. The camera system is automatic. It requires little space and is contained in one unit. The electronic still uses tubes. It lend itself excellent for lower resolution work or student training. If you need more info or any other help please contact me. Peter Jordan
Dear Coordinator, I write to confirm that I have received and read your mail concerning the Microscopy forumand that my particulars are correct. i will therefore be gratefull if you finally include me on your list.
I have had several years' experience of working with hardwood tissues in studying cambium and vascular differentiation (in Aesculus hippocastanum) for routine TEM and immunolocalization of cytoskeleton and cell wall and plasmalemma epitopes. If you have access to a library, can I suggest you look at Protoplasma vol 197, pages 64-75 (1997)(Chaffey et al. TEM, JIM-staining and cytoskeleton localization) and Int. J Plant Sci. vol. 158, pages 97-109 (1997) (Chaffey et al. TEM, Thiery staining, ZIO-staining and JIM-staining) for protocols and pictures? Also look at the two recent papers by Farrar and Evert in Trees vol. 11, pages 191-202 and 203-215 (1997) (so I don't just quote my own work!)
In short, for TEM I use 2.5% glut, 3.7% form in 25 mM PIPES buffer, pH 6.9, Os post-fix, alcohol dehydration, prop oxide transfer and Spurr resin. For JIM-staining, I use same primary fix, alcohol dehydration and LR white embedding either UV-cured at -20 C, or at c. 45 C. I have not yet got a TEM localization method for tubulin or F-actin in this system (although I can supply details of indirect immunolocalization of these components at the light level).
I hope some of the above is useful. If you want to know/ask more, please contact me direct (please supply fax number(s) if protocols required: I just can't get the hang of sending 'attachments'!).
Good luck!
Nigel Chaffey
----------------------------------------------------- Dr Nigel Chaffey, Dept Forest Genetics & Plant Physiology, Swedish University of Agricultural Sciences, SE-901 83 Umea, Sweden Phone: +46-90-166305 Fax: +46-90-165901 eMail: nigel.chaffey-at-genfys.slu.se
Excuse me about my short question. I believe that I have problems with the osmium because I haven't got that problem when the tissue is not osmicated. I buffered the osmium solution with phosphate and cacodylate buffer and the osmium precipitates appear in the sample (using different animals tissues, I haven't got the problem in plant cells). The protocol is as follow: .fixation: 0-3% paraformaldehyde and 1-2.5% glutaraldehyde in phosphate or cacodylate buffer 0.1M pH 7.4. Time: 30'- 12 h. Temp: Room temp. or 4C. .Rinse with the same buffer solution.Time: 4-5 x 10'. Temp: Room temp. .Postfixation: 1-2% osmium tetroxide buffered with the same buffer solution. Time: 1 h. 4C .Rinse with water (before the osmium precipitates problem we rinse with buffer). Time: 4 x 10'. Temp: room temp. .Dehydration : acetone .Embedding: Spurr, Araldite..
The last test is buffer the osmium solution with veronal acetate but I fhaven't got the results yet.
Thanks very much for your help
Nuria Cortadellas Department of Electron Microscopy University of Barcelona
I just read the FTIR postings and know nothing about this method. I note that they refer to particulates but can it be used to identify hydrocarbons? With pollution in mind. I remember years ago that I kept a leaflet about an instrument which could do this using very small samples (on microscope slides, if I remember correctly). Now, I can't find the leaflet!
I may have inadvertently sent two messages to the list that were intended for indivviduals. To compound the error they both have hefty (280k) attachments. Please accept my apologies.
On Wed, 11 Jun 1997 Robert414-at-aol.com wrote: About 5 years ago it was common for our Taiwanese students to have a locally printed book containing all five volumes of Edington. I understand that at that time in Taiwan the laws of copyright were different from in Britain.
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } I was wondering if anyone knows where I can get a copy of the Transmission } Electron Microscopy Monograph series by Edington. I believe they are out of } print but I was hoping that there may be a place that still has a supply of } them. Thanks. } Alasdair Preston Dept of Mat Sci and Met Cambridge University Pembroke Street Cambridge
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Reply to: RE} osmium precipitates
Dear Nuria, Sounds like your phosphate buffer interacting with the osmium may be the cause of precipitates. We generally fix for resin embedding in cacodylate buffer. If phosphate buffer is used for the fixative, we wash it away 3 x 10 minutes with cacodylate buffer then proceed with osmication. The osmium is also never diluted in phosphate buffer. We keep a stock solution of 4% made in water then dilute to 2% with cacodylate buffer when ready to use. Hope this helps. Linda Chicoine Center for Cell Imaging Dept. of Cell Biology Yale University 333 Cedar Street New Haven, CT 06520-8002 203-785-3646 tel. 203-785-7226 fax
--------------------------------------
Excuse me about my short question. I believe that I have problems with the osmium because I haven't got that problem when the tissue is not osmicated. I buffered the osmium solution with phosphate and cacodylate buffer and the osmium precipitates appear in the sample (using different animals tissues, I haven't got the problem in plant cells). The protocol is as follow: .fixation: 0-3% paraformaldehyde and 1-2.5% glutaraldehyde in phosphate or cacodylate buffer 0.1M pH 7.4. Time: 30'- 12 h. Temp: Room temp. or 4C. .Rinse with the same buffer solution.Time: 4-5 x 10'. Temp: Room temp. .Postfixation: 1-2% osmium tetroxide buffered with the same buffer solution. Time: 1 h. 4C .Rinse with water (before the osmium precipitates problem we rinse with buffer). Time: 4 x 10'. Temp: room temp. .Dehydration : acetone .Embedding: Spurr, Araldite..
The last test is buffer the osmium solution with veronal acetate but I fhaven't got the results yet.
Thanks very much for your help
Nuria Cortadellas Department of Electron Microscopy University of Barcelona
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A student asked me to query about the density of Fe3Si. I did not see it in the phase diagram available to me.
Thus does it exist and if it does do some have a measured density for it?
Thanks
## [########] ## ## ## ## ## Stephan H Coeztee Electron Microscope Unit Private Bag 3 Wits 2050 South Africa
I have always tried to avoid cacodylate buffer by using safer alternatives such as phosphate or Pipes but I am about to embark on some new work where all of the references have used cacodylate in the past. Obviously it is good practice to try and reduce the use of hazardous materials in the lab., especially in e.m., where we have so many. This is embodied in our safety laws in the UK especially in COSHH regulations (Control of Substances Hazardous to Health 1988) but, judging by recent worries on the list about formaldehyde and glutaraldehyde, is a principal that we all adhere to.
My question is: has anyone come up with a safer alternative to cacodylate that could be used as a direct substitute? Phosphate won't do because of its problems with Ca++ but Pipes seems safer (the safety data sheets I have are not as helpful as they could be), although it is expensive and probably won't keep as long.
A student from Botany is experiencing problems with staining cassava somaltic embryos at the globulas stage. She is embedding in Spurs Resin. Reynold's staining is used with the staining times: 30 min uranyl acetate 1 hr in lead staining
The staining is very week.
## [########] ## ## ## ## ## Stephan H Coeztee Electron Microscope Unit Private Bag 3 Wits 2050 South Africa
We have a DV-502 which we use mostly for TEM samples. In general it works just fine but you might want to keep in mind the following :
1)The carbon rod is spring loaded, and the spring that we purchased from Denton is quite stiff. This tends to cause a premature brake of the carbon tip. I would recomend changing this spring to one that is less stiff.
2) With time the bell jar and the various components within the chamber will get cabon coated and will have to be cleaned. Depending on the level of coating this can take quite some time. I would suggest that you modify the shield ( i.e. build a larger one ) that will result in less carbon build up in the surrounding areas and which will be easier to clean.
Jordi Marti.
P.S. Dending on the kind of work/samples a bench top model (evaporator and sputter unit) with a mechanical pump might be sufficient for your work and you might save some $$$. -----------------------------------------
Although Jan may know of some cache at PEO with these books (and different Philips personnel have different amounts remaining), my contacts with Philips over the past few years to obtain these volumes have indicated that the only remaining copies are the least requested volumes. The two (or three?) most used volumes have been out of stock for some time now. However, the volumes have been combined into a single volume and are still in print, or at least available, (although in a *much* poorer quality) from TechBooks. Techbooks is at 4012 Williamsburg CT Fairfax, VA 22032-1139 (703-352-0001). (I just found another Techbooks listing at: Campus Square Klamath Falls, OR 97601 (541-884-3882) but don't know if they would also have Edington's book.)
I have also seen a number of versions printed in Asia, but even aside from the copyright concerns, the quality of the ones I've seen has been no better (and may have been worse) than the TechBooks version.
Good luck in your search.
Dick Fonda
In message {39f71d60-at-pei.philips.com} jan ringnalda writes: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } The Edington books are still available through Philips Electron Optics } although supplies now are getting limited. } } Cheers, Jan
_____________________________________________________________ Richard W. Fonda Naval Research Laboratory (202) 767-2622 Code 6324 (202) 767-2623 fax Washington DC 20375 _____________________________________________________________
At 04:13 PM 6/11/97 EDT, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America It would all depend on the heat load being produced by the instruments that you are trying to cool. Check with your manufacturers. We once considered using a single Haskris with a "Y" in the supply line to serve two instruments with low heat load at the same time, while we were waitng for repair parts for the other Haskris. Hitachi service man though it should work. Ultimately we were able to get along without resorting to that, so I can't say for sure if it would work.
} Question #2: A client of mine wants to see if she can see the } difference between elastin and collagin at the TEM level {she is } looking for the relative amounts in various samples}. Is this } possible? What are the references/protocols? Am I nuts to try this?
Could antibodies be used to differentiate the two in some way } } Thanks, } John Grazul } Rutgers University } Electron Imaging Facility } } ******************************************************* 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
thanks to everyone who replied to my two very different questions. The collagen/elastin experiment will start next week if the researcher can get the samples, and the chiller will be retrofitted ASAP.
John Grazul Rutgers University Electron Imaging Facility
The purity of your glutaraldehyde may be the problem. At one time we re-distilled ours to avoid the "pepper" problem. We still sometimes have a the problem you describe especially in muscle. It appears from time to time for no explicable reason.
} } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } At 09:11 AM 6/12/97 +0200, you wrote: } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America 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
Here in Reading, we are still using a Philips 301 TEM with a 35 mm camera. Until about a year ago, we were using Agfa Scientia 35 mm unperforated roll, when this was discontinued. Many people have offered us Eastman 5302, but this has two disadvantages (1) it is only about 25% as sensitive to electrons (2) it suffers stress marks when packed too tightly into the cassette.
We have already contacted microscopy groups on an individual basis, from Finland in the North to New Zealand in the South, and it appears this situation is global. If you have any ideas, please contact us IF:
(a) you know of another source of film; (b) you would like to see 35 mm Agfa Scientia, and would like to form a group to approach the company.
Please don't reply simply to say "we use plates", on the principle:
"I only want a little bit of butter for my bread" "But many people nowadays like marmalade instead"
(from A.A.Milne)
Thank you
+------------------------------------------------------------------------+ | Robert H.Olley Phone: | | J.J.Thomson Physical Laboratory {direct line +44 (0) 118 9318572 | | University of Reading {University internal extension 7867 | | Whiteknights Fax +44 (0) 118 9750203 | | Reading RG6 6AF Email: R.H.Olley-at-reading.ac.uk | | England URL: http://www.reading.ac.uk/~spsolley | +------------------------------------------------------------------------+
I do not like Spurr Resin because it gives inconsistent results. Sometimes it stains poorly. Try Epon 812 or its substitutes or LR White. Lead staining for 1 hr seems too long and it is likely to precipitate. I usually stain for 5 min. on Epon sections.
A student from Botany is experiencing problems with staining cassava somaltic embryos at the globulas stage. She is embedding in Spurs Resin. Reynold's staining is used with the staining times: 30 min uranyl acetate 1 hr in lead staining
The staining is very week.
Stephan H Coeztee Electron Microscope Unit Private Bag 3 Wits 2050 South Africa
I saw your posting on the listserver and just wanted to re-send my message dated 6/5/97 in case that you did not receive it.
Dear Richard:
Since you are getting faint lattice images from the YBCO sample that we prepared, it is quite possible that the carbon film is the source of scattering. The "conventional" method of FIB preparation requires that the specimen is mounted on a grid, but a carbon support is not needed, that is, the electron beam will be passing through the electron transparent area only. We can try to prepare the YBCO film using conventional methods of FIB specimen preparation. We might be able to try it with the remaining sample, but this technique might require a larger piece of sample. Can you send us an additional piece?
Can you also please forward the lattice parameters of YBCO to me so that we can test and view the specimen for lattice images prior to shipping it to you?
If this specimen works out, we will charge you the previously quoted rates for TEM FIB specimen preparation:
TEM Sample Preparation:
FIB sample preparation $250/hr other sample preparation $150/hr
TEM work $200/hr
I anticipate that the preliminary specimen preparation might require up to 5 hours, FIB usage will probably be about 3 hours with no more than 5 hours of use. The TEM will be used for ~ 1 hour to check for the usefulness of the specimen. Based on these figures, the specimen can be prepared for ~ $1,700 - $2,200. If this is something that you would like to pursue please let me know at your earliest convenience.
Thank you,
Lucille Giannuzzi
************************************************************************* Lucille A. Giannuzzi, Ph.D. Assistant Professor
Dept. of Mechanical, Materials, and Aerospace Eng.
University of Central Florida phone (407) 823-5770 PO Box 162450 fax (407) 823-0208 4000 Central Florida Blvd. email lag-at-pegasus.cc.ucf.edu Orlando, FL 32816-2450 USA *************************************************************************
Dear Microscopists, Our laboratory urgently needs formula for preparation Lowicryl K4M resin for low temperature embedding after freeze-drying. We have all set of chemicals in hand, but without any precise instruction how to mix them. Thanks in advance for all help Best regards Jolanta Przybylowicz
Independent from the fixative I would consider using Lowicryl HM20 instead of LR White, its viscosity is much lower and immuno labelling works nicely. For example aleurone cells of barley grains (which have huge cell walls, which even represent a problem with Spurrs) are easy to infiltrate with this stuff at -35 C, and organells are preserved quite well.
Good luck, 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
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } I will be involved with operating an ESEM in the near future and need } information about any courses available concerning this technology. Any } information will be greatly appreciated. } } Well Gary, I don't know where you are at Geographically, - But if anyone is interested - The Institute of Meteoritics at the University of New Mexico holds an EM course once a year that includes teaching on the EMP, SEM, ESEM, and LVSEM, as well as sample prep, limitations, new technology and an overview of Secondary Ion Mass Spec. (SIMS). We also teach courses in TEM, ICPMS, and a number of other types of equipment (XRD, OM...etc...). Of course the problem is that you have to get to New Mexico to take the course.
For More Info, reply to yoyodine-at-unm.edu
I would suggest to anyone looking for courses in Microscopy of any kind to contact the nearest University...classes of this type or offered in depts. such as Materials Science, Earth and Planetary Science, Geology, Biology, Chemistry, and Engineering...SEMS are rather common and ESEM are basically the same machine with the addition of a gated vacuum system...any hands on SEM course should do the trick.
Hope this helped Christopher Adcock
PS: I replied to all on the list because I thought others might be interested, but I have noticed that I get alot of clutter in my mbox and others most likly do to. So if you want more info, it would be best to reply to me alone and keep others boxes clean.
In some cases, cacodylate is the preferred buffer because it helps the fixative to penetrate the tissue faster and further. Certainly, it is hazardous, but if one uses a fume hood and wears gloves one can minimise the danger, and of course you should be doing this anyway, since the fixative itself is so much more dangerous than the cacodylate.
Lesley Weston
On Thu, 12 Jun 1997, HASWELL Malcolm wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } Dear all } } I have always tried to avoid cacodylate buffer by using safer alternatives } such as phosphate or Pipes but I am about to embark on some new work where } all of the references have used cacodylate in the past. Obviously it is good } practice to try and reduce the use of hazardous materials in the lab., } especially in e.m., where we have so many. This is embodied in our safety } laws in the UK especially in COSHH regulations (Control of Substances } Hazardous to Health 1988) but, judging by recent worries on the list about } formaldehyde and glutaraldehyde, is a principal that we all adhere to. } } My question is: } has anyone come up with a safer alternative to cacodylate that could be used } as a direct substitute? Phosphate won't do because of its problems with Ca++ } but Pipes seems safer (the safety data sheets I have are not as helpful as } they could be), although it is expensive and probably won't keep as long. } } thanks in advance, } } Malcolm Haswell } University of Sunderland } UK } } }
} Excuse me about my short question. I believe that I have problems with } the osmium because I haven't got that problem when the tissue is not } osmicated. I buffered the osmium solution with phosphate and cacodylate } buffer and the osmium precipitates appear in the sample (using different } animals tissues, I haven't got the problem in plant cells). The protocol } is as follow: } .fixation: 0-3% paraformaldehyde and 1-2.5% glutaraldehyde in phosphate or } cacodylate buffer 0.1M pH 7.4. Time: 30'- 12 h. Temp: Room temp. or 4C. } .Rinse with the same buffer solution.Time: 4-5 x 10'. Temp: Room temp. } .Postfixation: 1-2% osmium tetroxide buffered with the same buffer } solution. Time: 1 h. 4C } .Rinse with water (before the osmium precipitates problem we rinse with } buffer). Time: 4 x 10'. Temp: room temp. } .Dehydration : acetone } .Embedding: Spurr, Araldite.. } } The last test is buffer the osmium solution with veronal acetate but I } fhaven't got the results yet.
If freshly depolymerized paraformaldehyde is not used, the short formaldehyde 'mers are left behind in your tissues and react with the osmium to form granular precipitates (very small, peppery looking).
Phosphate buffers are notorious precipitators (especially with uranyl salts) but cacodylate should be precipitate free.
In some (perhaps a lot of) instances it is not necessary to buffer the osmium - simply use an aqueous solution.
Is your distilled water good? Water containing volatile amines or other organics may react with the osmium/aldehydes to cause precipitates.
Please describe the precipitates (large, anglular, blobs, everywhere, localized, etc).
#################################################################### 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 trying to locate a laboratory or University that has Scanning White Light Interferometry available for contract research. The two instruments I am familar with and have provided us with useful data are manufactrued by Zygo and WYKO.
Thanks, John Catino Union Camp Corp. Princeton, NJ
Absolutely - Actually, I think I can speak for the other respondents in saying that most of the particles and fibers that we are referring to are organic in nature. Organic compounds (i.e. various forms of hydrocarbons) are the primary analytes examined by FTIR. Generally speaking, depending upon the sample condition and matrix, one can obtain very good qualitative and even quantitative FTIR assays. CH absorptions abound in the following regions of the spectrum (in wavenumber):
2800-3000 cm-1 1300-1490 cm-1
If you were to collect airborne hydrocarbon particulates onto filters, you could qualitatively analyze these by micro-FTIR - you may not get a positive identification of a single compound (because you are likely to have a mixture), but you might be able to identify chemical families.
Regards,
Bob ************************* Bob Citron Chiron Vision Claremont, CA USA (909)399-1311 Bob_Citron-at-cc.chiron.com **************************
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Hello from Plymouth, UK
I just read the FTIR postings and know nothing about this method. I note that they refer to particulates but can it be used to identify hydrocarbons? With pollution in mind. I remember years ago that I kept a leaflet about an instrument which could do this using very small samples (on microscope slides, if I remember correctly). Now, I can't find the leaflet!
Just want to thank all of you for your responses to my question on xylene replacements. I now have a number of leads which I will follow up on shortly.
Paula.
Paula Allan-Wojtas Food Microstructure Specialist Agriculture and Agri-Food Canada Atlantic Food and Horticulture Research Centre Kentville, Nova Scotia Canada B4N 1J5
To get good electron channeling patterns you need a surface that is completely free of any form of mechanical distortion such as can be introduced by most mechanical cutting, grinding and polishing operations. If you must use mechanical techniques to prepare your specimen's surface, then you must etch it rather heavily after each stage(after cutting, after grinding, and after each polishing step) to remove as much cold-worked metal as possible, and at the end you must use a much heavier etch than you normally would for just microscopic analysis - you must see clearly defined, distortion-free grains. This will require some experimentation to find an etchant that will remove lots of metal without pitting. If possible, it is preferable to use electrolytic polishing, because this does not introduce mechanical deformation. To get started, might I suggest getting a piece of single crystal silicon of the type used in the semiconductor industry, which should give good channeling patterns as-received, and play around with that until you get the hang of the technique.
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 have found dimethylsulfoxide (DMSO) to be a truly remarkable material to use for treating minor burns of the ordinary variety. I have found all the claims made on p. 107, and elsewhere, in the book 'DMSO, The Pain Killer' (by Barry Tarshis, Berkley Books, 1981, ISBN: 0-425-07488-9) to be unbelievably true. The minute you experience a common-type, 'small' burn, wash it for a few minutes with cold water, then apply a 70% DMSO-30% water solution libally immediately, and then two or three more times at about one-hour intervals, and then again twice daily for a couple more days. I find this procedure will usually stop the pain almost immediately, and more surprisingly, will usually keep the burn from blistering. I have found it also works if you begin to develop a blister from having a shoe rub your heel, or from raking the lawn without gloves, etc.- prompt and persistent application of DMSO will usually relieve the pain and keep a blister from developing. I haven't tried it for LN2 burns, but would sure give it a try if I happened to experience one. I keep a bottle in my office, and another at home, for just such occurrences.
Dimethlysulfoxide is not a material that is approved for medical use, and so if you make use of it you do so under your own responsibility; however, it has long been used by athletes to relieve the pain from bruises and sore muscles, and it is reported (see above source) to also be effective in relieving the pain associated with some cases of bursitis, arthritis, interstitial cystitis, scleroderma, headaches, gout,hemorroids, herpes infections, shingles, etc., etc.
DMSO is not medically approved because it is a cheap by-product of the pulp and paper industry, and therefore not patentable as a drug. Thus, no drug company is willing to go to the expense of running the necessary test to get it approved. In addition, most people develop a garlic-like taste from using it, and so it is almost impossible to run the double-blind tests needed to gain such approval. It is, however, widely available in health food stores.
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
Many thanks for your kind input regarding my PBS argument. I have decided to use a slightly hypertonic (4mM) phosphate buffering system in 9% saline solution. The slight hypertonicity (and slightly increased buffering capacity) may help preserve tissue during the DAB reaction, because this tissue still has semi-permeable membranes due to the absence of osmium fixation. Thanks again. This Listserver is better than chocolate! Bye, Hildy
john catino wrote: } } ------------------------------------------------------------------------} The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------.} } I am trying to locate a laboratory or University that has Scanning White } Light Interferometry available for contract research. The two } instruments I am familar with and have provided us with useful data are } manufactrued by Zygo and WYKO. } } Thanks, } John Catino } Union Camp Corp. } Princeton, NJ } } -- } II*Dear John,
Both Zygo and Wyko have facilities here in the Northeast which are available on a contract basis. Call Zygo at (460)247-8372 (Ask for Barbara - not me) and Wyko at (602)741-1297.
} } P.S. There was a thread awhile ago on what gloves to use with what } } chemicals--was there a definitive list that came out of that discussion? I } } know glut goes through latex like the glove isn't there. P } } } } } Sic Hoc Legere Scis Nimium Eruditionis Habes { } } Philip Oshel
Phil and anyone else who is interested, I spent some time researching the issue of what gloves our Unit should be recommending for all our users. As a result I drew up a table of which gloves to wear when handling various common chemicals in our Unit. I can send anyone interested a copy of this, however I take no responsibility for the information contained in it in any way whatsoever (except to people using our EM Unit), I'm still updating it as I learn more.
The whole subject is more complex than I first thought. There is quite a bit of apparently contradictory information on glove suitabilty, largely resulting from the large number of different glove polymers, glove styles, chemical combinations and work practices around. Because we are a multiuser lab with about 90 users on our books at present we really were only interested in disposable gloves. Thick, heavy gloves that hindered fine manipulation were also out. I came to the conclusion that, with a few important exceptions, latex gloves are actually fairly good for most of the chemicals we used based on the studies I read.
Although latex has a poor reputation for use with glutaraldehyde there is at least one paper (Jordan et al, Glutaraldehyde permeation: Choosing the right glove, Union Carbide 1996) which guardedly suggests that it is probably OK for use with the low concentrations of glutaraldehyde usually used for fixing tissue, as long as contaminated gloves are removed as soon as possible. That really is one of the crucial aspects of wearing gloves for chemical protection - all gloves are permeable eventually to many chemicals and one of the best ways to minimise exposure risk is to remove gloves as soon as possible after contamination and don new ones. In the above paper the authors state that the breakthrough time for a single layer of latex examination gloves exposed to 2% glutaraldehyde was between 30 and 45 minutes (at 25degC however). This is possibly good enough for people dealing with specimens in low concentrations of glutaraldehyde, provided they don't continue to wear the gloves when contaminated and provided the individual is not sensitised to glutaraldehyde. Therefore it is probably not true to say that 'glut goes through latex like the glove isn't there', it does provide short-term protection. Whether it provides enough is another matter, it may be possible that latex permits enough glutaraldehyde through to eventually cause sensitisation to glutaraldehyde. If you want to be safer, nitrile and butyl synthetic rubber provide a much better barrier than latex.
I think that if you want to minimise the risk you should use nitrile instead of or as well as (over) latex for glutaraldehyde. I personally get slightly itchy hands from using our disposable nitrile gloves so I tend to wear them over latex gloves. Nitrile is also supposed to be better as a barrier for acrylic resins too. You can't substitute disposable nitrile gloves for latex totally however because nitrile doesn't cope with exposure to some resins and solvents. Nitrile is particularly ineffective as a barrier against propylene oxide (less effective than latex anyway - however latex isn't too good against propylene oxide either).
You can apparently get extra protection by using one of the spray-on barrier creams too.
Regards,
Richard
Richard Easingwood South Campus Electron Microscope Unit School of Medical Sciences University of Otago PO Box 913 Dunedin NEW ZEALAND
Subject: Time:4:25 PM OFFICE MEMO Need EM200 sample holder Date:6/12/97
Hello All,
We're looking for an inexpensive TEM sample holder for the Philips EM200 or EM 400 series microscopes. Used, simple, single tilt, is fine, if not bent. Key parameter is inexpensive.
DGCollins-at-lbl.gov (510) 486-7859, or DCollins 2841 Kinney Dr, Walnut Creek, CA 94595 (510)939-2006
*Received: from SpoolDir by IM-PW (Mercury 1.13); Thu, 12 Jun 97 23:33:05 -1800 *Return-path: {Microscopy-request-at-sparc5.microscopy.com} *Received: from Sparc5.Microscopy.Com by mailer.inmat.pw.edu.pl (Mercury 1.13); * Thu, 12 Jun 97 23:32:54 -1800 *Received: (from daemon-at-localhost) by Sparc5.Microscopy.Com (8.6.11/8.6.11) id PAA06007 for dist-Microscopy; Thu, 12 Jun 1997 15:00:13 -0500 *Received: from srvr7.engin.umich.edu (srvr7.engin.umich.edu [141.212.2.69]) by Sparc5.Microscopy.Com (8.6.11/8.6.11) with ESMTP id PAA06004 for {microscopy-at-sparc5.microscopy.com} ; Thu, 12 Jun 1997 1
*:00:12 -0500 *Received: from [141.212.131.74] (dow2146amac74.engin.umich.edu [141.212.131.74]) * by srvr7.engin.umich.edu (8.8.4/8.8.4) with ESMTP * id QAA25518 for {microscopy-at-msa.microscopy.com} ; Thu, 12 Jun 1997 16:00:00 -0400 (EDT) *X-Sender: bigelow-at-srvr5.engin.umich.edu *Message-Id: {v03007802afc5bce8e0dc-at-[141.212.131.74]} *Mime-Version: 1.0 *Content-Type: text/plain; charset="us-ascii" *Date: Thu, 12 Jun 1997 10:58:02 -0400 *To: Microscopy Listserver {microscopy-at-Sparc5.Microscopy.Com} *From: Wil Bigelow {bigelow-at-engin.umich.edu} *Subject: RE:Electron Channeling
Professor is absolutly right. There should be also kept in mind that the investigated material should be free of cold work in order to obtained sharp channeling patern. In the past, there was done a lot of work by Prof. W.W.Gerberich and members of his research group on influance of strain on channeling pattern.
Witold Zielinski Warsaw University of Technology Narbutta 85 02-524 Warszawa, Polska
I am interested in gloves resistant particularly to acetone, which most of our dehydration is done in.
Having seen the initial posting, I tried a crude test today on some new, thin nitrile gloves: 15 minutes dangling the finger ends (empty!) in conc. HNO3, 25% glut and pure acetone (in separate beakers). Then the gloves were washed carefully on their exterior and dabbed dry with a towel. The HNO3 trial showed a serious failure problem! The other gloves were then blown up by mouth and tested with the Mark I nose. The acetone penetration was serious, but the glut was not detectable (and this Mark I nose is normally quite sensitive). That's it.
} From: "Robert H. Olley" {R.H.Olley-at-reading.ac.uk} } To: Microscopy Newsgroup {Microscopy-at-Sparc5.Microscopy.Com} } Cc: # {R.H.Olley-at-reading.ac.uk} } Subject: TEM: 35 mm film - where to find? } } Here in Reading, we are still using a Philips 301 TEM with a 35 mm } camera. } Until about a year ago, we were using Agfa Scientia 35 mm unperforated roll, } when this was discontinued. Many people have offered us Eastman 5302, but } this has two disadvantages (1) it is only about 25% as sensitive to } electrons (2) it suffers stress marks when packed too tightly into the } cassette. } } We have already contacted microscopy groups on an individual basis, from } Finland in the North to New Zealand in the South, and it appears this } situation is global. If you have any ideas, please contact us IF: } } (a) you know of another source of film; } (b) you would like to see 35 mm Agfa Scientia, and would like to form a } group to approach the company. } Thank you } } +------------------------------------------------------------------------+ } | Robert H.Olley Phone: | } | J.J.Thomson Physical Laboratory {direct line +44 (0) 118 9318572 | } | University of Reading {University internal extension 7867 | } | Whiteknights Fax +44 (0) 118 9750203 | } | Reading RG6 6AF Email: R.H.Olley-at-reading.ac.uk | } | England URL: http://www.reading.ac.uk/~spsolley | } +------------------------------------------------------------------------+ } Dear Mr. Olley,
Many of our M. users have already changed to Kodak 35 mm Technical Pan Film and are very positive about it. However with respect to Agfa Scientia and Kodak 5302 it has two disadvantages: it is perforated and you need a special developer.
Regards,
Bert Boxstart Philips Electron Optics B.V. Comm. Support dep.
Hi I am urgently looking for the E-mail address of either Dr DJ Cork or J P Krueger from the Illinois Institute of Technology in Chicago. If you can be of assistance please reply to my E-mail address.
I apologize for this non-related microscopy request
Thank you Alan N Hall Unit for Electron Microscopy Faculty of Biological & Agricultural Sciences University of Pretoria Pretoria 0002 Republic of South Africa Tel: +27-12-420 3297 Fax: +27-12-420 3266
Sorry this is so late in the train of information here, but I have been told by two or three technicians who use DAB regularly to use Tris buffer instead of phosphate buffer with the DAB solution. One even had me doing a Tris buffer rinse after a fix with PBS to make sure there was no phosphate buffer remaining at the DAB-labeling step. I forgot what the problem was, but maybe someone else out there can add some info. The point is, you may have solved the tonicity problem, but you could still have some problem with the DAB substrate reaction if you are using PBS with the DAB. P.S. I got the recipe for Tris buffer from Handbook of Immunoperoxidase Staining Methods by Janice A. Bourne, published by DAKO Corp. in 1983. Tris buffer (0.05M) is 6.1g trishydroxymethyl aminomethane (tris base), 50 ml dH2O, mix and add 37 ml 1N HCl, dilute to 1 liter with dH2O. The pH should be 7.6+/- 0.2 at 25oC (I often use a pH or 7.2, just adjust the volume of HCl). They also have a recipe for Tris buffered saline using 100ml of this Tris with 900ml of 0.85% saline or PBS. They say it can reduce background staining.
Another way to help preserv tissue at the DAB step is to include a non-peroxide DAB step prior to the actual reacting step. I find that this enhances the reaction and often cuts down the time of exposure to the peroxide. Hope this is helpful.
Karen
On Thu, 12 Jun 1997, HILDEGARD CROWLEY wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } } Dear Folks, } } Many thanks for your kind input regarding my PBS argument. I have } decided to use a slightly hypertonic (4mM) phosphate buffering system in } 9% saline solution. The slight hypertonicity (and slightly increased } buffering capacity) may help preserve tissue } during the DAB reaction, because this tissue still has semi-permeable } membranes due to the absence of osmium fixation. } Thanks again. This Listserver is better than chocolate! } Bye, } Hildy }
I am working with the human cornea and am exploring options for tissue preparation techniques and staining procedures for LM. Sections of 0.5-1.0 microns will be used to distinguish between the epithelium and underlying Bowman's layer. I need to find the optimal embedding medium as well as staining procedure.
I'd like to know how could I get histological specimens from titanium implants inserted into rabbit bone. The problem is cutting bone with metal. It seems that the suitable equipment is named Exact. How does it work? What about its price? Where can I get it?
Yours sincerly,
Marcelo Henrique Prado PEMM - COPPE/UFRJ Po.Box.:68505 Cidade Universitaria - Ilha do Fundao Rio de Janeiro-R.J. CEP.: 21941-900
Richard Lander wrote: } } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Phil Oshel wrote; } } } } P.S. There was a thread awhile ago on what gloves to use with what } } } chemicals--was there a definitive list that came out of that discussion? I } } } know glut goes through latex like the glove isn't there. P } } } } } } } Sic Hoc Legere Scis Nimium Eruditionis Habes { } } } Philip Oshel } } Phil and anyone else who is interested, } I spent some time researching the issue of what gloves our Unit should be } recommending for all our users. As a result I drew up a table of which } gloves to wear when handling various common chemicals in our Unit. I can } send anyone interested a copy of this, however I take no responsibility for } the information contained in it in any way whatsoever (except to people } using our EM Unit), I'm still updating it as I learn more. } } The whole subject is more complex than I first thought. There is quite a } bit of apparently contradictory information on glove suitabilty, largely } resulting from the large number of different glove polymers, glove styles, } chemical combinations and work practices around. } Because we are a multiuser lab with about 90 users on our books at present } we really were only interested in disposable gloves. Thick, heavy gloves } that hindered fine manipulation were also out. I came to the conclusion } that, with a few important exceptions, latex gloves are actually fairly } good for most of the chemicals we used based on the studies I read. } } Although latex has a poor reputation for use with glutaraldehyde there is } at least one paper (Jordan et al, Glutaraldehyde permeation: Choosing the } right glove, Union Carbide 1996) which guardedly suggests that it is } probably OK for use with the low concentrations of glutaraldehyde usually } used for fixing tissue, as long as contaminated gloves are removed as soon } as possible. That really is one of the crucial aspects of wearing gloves } for chemical protection - all gloves are permeable eventually to many } chemicals and one of the best ways to minimise exposure risk is to remove } gloves as soon as possible after contamination and don new ones. In the } above paper the authors state that the breakthrough time for a single layer } of latex examination gloves exposed to 2% glutaraldehyde was between 30 and } 45 minutes (at 25degC however). This is possibly good enough for people } dealing with specimens in low concentrations of glutaraldehyde, provided } they don't continue to wear the gloves when contaminated and provided the } individual is not sensitised to glutaraldehyde. } Therefore it is probably not true to say that 'glut goes through latex } like the glove isn't there', it does provide short-term protection. Whether } it provides enough is another matter, it may be possible that latex permits } enough glutaraldehyde through to eventually cause sensitisation to } glutaraldehyde. If you want to be safer, nitrile and butyl synthetic } rubber provide a much better barrier than latex. } } I think that if you want to minimise the risk you should use nitrile } instead of or as well as (over) latex for glutaraldehyde. I personally get } slightly itchy hands from using our disposable nitrile gloves so I tend to } wear them over latex gloves. Nitrile is also supposed to be better as a } barrier for acrylic resins too. You can't substitute disposable nitrile } gloves for latex totally however because nitrile doesn't cope with exposure } to some resins and solvents. Nitrile is particularly ineffective as a } barrier against propylene oxide (less effective than latex anyway - however } latex isn't too good against propylene oxide either). } } You can apparently get extra protection by using one of the spray-on } barrier creams too. } } 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 think it should be clearly stated that neither latex nor nitrile gloves provide any effective barrier to epoxy and acrylic resins. I became sensitized, and had a very severe reaction to acrylic resins through the misconception that nitrile gloves were the gloves of choice for handling resins. The only glove I would recommend to anyone working with resins is the 4H glove from Safety 4 A/S, 9765 Widmer, Lenexa KS 66215 (913) 492 0860. Some people may find them a bit cumbersome at first, but we usually put a pair of nitrile gloves over the 4H gloves (not for added protection, but for a better finger feel). Don't forget to also wear a protective sleeve (also 4H), as the area between the glove and lab coat is still susceptible to the fumes of the monomer. The 4H glove also has a break-through time of } 240 minutes with a 25% glutaraldehyde solution. This is my personal opinion, after much research, and suffering, and does not represent the views of Texas Scottish Rite Hospital for Children.
Ronnie Houston Histology Coordinator Texas Scottish Rite Hospital for Children Dallas, TX 75219
On Thu, 12 Jun 1997 MESJASZ-at-NACDH4.NAC.AC.ZA wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Our laboratory urgently needs formula for preparation Lowicryl K4M resin } for low temperature embedding after freeze-drying. We have all set of } chemicals in hand, but without any precise instruction how to mix them. } Thanks in advance for all help
Hi Jolanta,
Here is the K4M recipe:
Cross linked A 2.7 g Monomer B 17.02 g Initiator C 20.1 g
The dehydration can be done with a series of ethanol at a low temperature. For the infiltration, the following schedule can be carried out at a low temperature.
1 : 1 ethanol : embedding medium 1 hr 1 : 2 ethanol : embedding medium 1 hr 100 % embedding medium, 2 X 1 hr, ea.
polymerization at -30 to -40 C for 24 hrs.
Good luck
*********************************************** * Ming H. Chen, PhD * * Medicine/Dentistry Electron Microscopy Unit * * University Of Alberta. * * Edmonton, Alberta, Canada * * * * Visit My Page At: * * http://www.ualberta.ca/~mingchen * ***********************************************
------------------------------------------------------------------------ The Microscopy ListServer -- Sponsor: The Microscopy Society of America
Subject: Time:7:32 AM OFFICE MEMO Post-doc position at LLNL Date:6/11/97
Our Center for Accelerator Mass Spectrometry (CAMS) is currently seeking a Postdoctoral Research Staff Member to work as part of an interdisciplinary team of physicists, chemists, and biologists. In this research position, you will work as part of a team to develop quantitative elemental analysis of biological tissues. These techniques will be used in studies of elemental kinetics, structural biology, toxicology or pharmacology by staff at LLNL and collaborators in the University of California system and other institutions. Duties include development/modification of techniques for preparation of biological tissues for quantitative elemental microanalysis, operation of the LLNL microprobes to analyze prepared biological samples, X#030#ray data reduction and analysis, planning/designing and executing independent and collaborative research projects and publication of results in peer#030#reviewed literature. Requires a recent Ph.D. in chemistry, biochemistry, toxicology, pharmacology or related field. Experience in trace element analysis and analytical microscopy is desired. LLNL offers a challenging work environment and a competitive salary/benefits package. Qualified individuals are invited to send their resume to: Lawrence Livermore National Laboratory, Attn.: Mary Anne Holman, L#030#725, P.O. Box 5510, R#030#4249, Dept. AJSC527MH, Livermore, CA 94551#030#5510. LLNL is an Equal Opportunity Employer. Lawrence Livermore National Laboratory Or contact me Graham bench Ph 510-423-5155 Cheers Graham Bench
You have unfortunately, just missed the best course on the subject. The Lehigh Short Course on SEM and Microanalysis. Contact Sharon Coe at {slc6-at-Lehigh.edu} for details od next years course.
Patrick Echlin
On Wed, 11 Jun 1997, Gary Lovell wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } I will be involved with operating an ESEM in the near future and need } information about any courses available concerning this technology. Any } information will be greatly appreciated. } }
I cannot tell you how much I appreciate and enjoy your replies to my problems. Our department is rather isolated here away from the town's huge medical research center, so instead of working with a vaccum, we often work in a vaccum! I am unable to repay you all individually, but I will take the time and effort to answer any problems on the LIST SERVER on which I feel I have expert information, and in such manner hope to be helpful to some of you who have gotten me out of a corner. (Corners) Thanks again! Hildy Crowley University of Denver Denver, CO
} DMSO is not medically approved because it is a cheap by-product of the } pulp and paper industry
In a chemical lab, DMSO is a dangerous substance because it takes any chemicals you might have on your skin and carries them into your blood. Anyone who applies DMSO to their skin should do so only after washing thoroughly and then not use any chemicals until the DMSO is really gone. Also the bottle of DMSO should not be kept near any toxic chemicals, and you should be sure that no one else has contaminated it with toxic chemicals. If I were using DMSO (I am not), I would want to use a new sealed bottle each time. What if the soap contains something toxic? What if the soap fails to remove something toxic?
} Dimethlysulfoxide is not a material that is approved for medical use, and } so if you make use of it you do so under your own responsibility; however, } it has long been used by athletes to relieve the pain from bruises and sore } muscles, and it is reported (see above source) to also be effective in } relieving the pain associated with some cases of bursitis, arthritis, } interstitial cystitis, scleroderma, headaches, gout,hemorroids, herpes } infections, shingles, etc., etc.
I have found Bigelow's information interesting but it left out a very important detail about DMSO's use by athletes, especially male athletes. DMSO has the ability to neutralize testosterone. Many male athletes thrive on testosterone as a feature needed for competition. This is one of the reasons that athletic trainers quit using DMSO.
Another problem with topical application of DMSO is that it can take with it live biological materials that may be on the surface of the skin. So it can pull with it virus for example that would not normally gain access to the interior of the body. For these and other reasons DMSO has not been approved for "general" medical use.
As I read Bigelow's last sentence above, snake oil comes to mind.
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
Gustave H. Wanner wrote: ---------------------------------------------------------------------- } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Co ---------------------------------------------------------------------- } Hi Everyone, } } Do any of you folks have any experience using the Polaroid MicroCam } with a Nikon Labophot-2 or equivalent microscope with Color-Free } objectives? The MicroCam replaces the eyepiece of the microscope; } it is unclear to me whether or not the MicroCam includes compensation for lateral chromatic aberation (also known as chromatic difference of magnification). The Nikon (as well as other recent microscope) objectives are corrected to provide a color free intermediate image. If the MicroCam includes compensation, it seems to me that this might negatively affect the resulting photomicrographs. } I have been unable to obtain any information from Polaroid telephone technical support on this question. } } Best regards, } } Gus Wanner } Exitech Corporation } 102 E. Broadway } Maryville, TN 37804 } (423) 983-9101 ListServer-at-MSA.Microscopy.Com Gustave H. Wanner {ghw-at-EXITECH.com}
Hello Gus, I am retired now from Polaroid, but was the microscopist on the development team for the MicroCam. I investigated just the question that you raised concerning Chromatic Difference of Magnification. I tested the MicroCam on a Nikon Optiphot with F head, using a 10X/0.25 CF objective, and a stage micrometer as the target, and found no color fringing.
I tested for fringing by focusing on a stage micrometer, or on the fine grating area of an Air Force Resolution Test Target; both provided a flat, high contrast specimen. I then looked for color fringing (orange or blue) in the image near the edge of the micrograph.
In making the MicroCam, it was our goal to make an inexpensive, ($795) camera compatible with a large number of microscopes, even those without phototubes, providing through the lens viewing, and automatic exposure control, --- and ease of use. These goals led to using an eyepiece integral to the camera.
As you mentioned, the lateral chromatic abberation, also known as CDM, for Chromatic Difference of Magnification, affects the compatibility of eyepieces with objectives. CDM is the difference in magnification between red and blue images at the primary image plane. CDM occurs in the objective and is the result of correcting for axial chromatic abberation in apochromatic objectives. This difference has traditionally been corrected in the eyepiece, and a manufacturer will typically design the same amount of correction into all their objectives so that all can be used with the same eyepiece. This practice has led to the advice against mixing eyepieces and objectives from different manufacturers.
(The Handbook of Optics, published by the Optical Society of America and McGraw Hill lists a number of eyepieces from a variety of manufacturers. The CDM for many of these eyepieces were listed, with all in the range of +0.3% to -1.5%. The microscopes in my laboratory used eyepieces which had a -1.4% correction, and the chromatic abberations were discernible at the periphery of the micrographs produced on those scopes if a correcting lens was not used.
Nikon introduced their "Chrome-Free" optics in 1976, and the Zeiss microscopes introduced in 1985 also corrected for CDM in the intermediate optics of the microscope. The objectives used in stereo microscopes do not exhibit CDM and do not need CDM corrections in the eyepiece. (Wild microscopes are an exception.)
Since one of the major uses for the MicroCam is for stereo microscopes, (often having no other photographic capability), and since the trend was toward correction before the eyepiece, we chose to incorporate in the MicroCam an eyepiece with no CDM. Therefore no fringing should be seen with chrome free optics, and with most stereo microscopes. (Some zoom systems do introduce some fringing.)
One response comment that you received had a number of complaints about the MicroCam system. The first concerned focus and resolution. Care with focusing is of course critical. It is important that the focusing crosshair be seen sharply before focusing the image. The MicroCam uses integral film, so exposure of the negative is through a clear polyester cover sheet. This makes the integral film sensitive to excess flare, so it is also important the substage iris of the microscope be adjusted to minimize flare and maximize contrast. (There is always the balance between contrast and resolution, and the condenser iris should be set at that point just before resolution is lost.) This comment may seem pedantic, and you might respond "of course", but the control of flare is particularly important with this film. The resolution of the color integral film, is ~ 10 line pairs per millimeter. This matches the resolution in the microscope image as governed by the resolution capabilities of most microscope objectives. Integral black and white films have a higher resolution, ~20 line pair per millimeter.
The second concerned color rendition. I have a number of commercially prepared, moderately stained slides, mainly H&E, some aniline blue and I have been satisfied with their color rendition. The MicroCam incorporates a light balancing filter approximating an 80B, which would be the appropriate filter for a tungsten halogen lamp at its rated voltage. Some adjustment of filtration may be necessary with the users own filters, to optimize color rendition for their light sources or for transmission characteristics of their microscopes.
Third, indeed, you do wind up with a print, and not a negative or a transparency. The print is self-developing outside the camera. I have used it with a Polaroid scanner, to get an electronic image for image analysis or incorporation in communications.
The fourth point mentioned that "Exposure varies with the specimen so many test shots are necessary." The exposure control measures the entire image area, and responds similarly to any other automatic camera system averaging a large area. The camera has electronic correction for reciprocity failure, both for speed and for color balance. The exposure adjustments are clear and easily accessible, for specimens with widely varying backgrounds.
I hope I have answered your question, and some of the other questions which have been raised. If I can be of further help, my email address is "mccanns-at-tiac.net".
Having retired from Polaroid, I no longer have any financial interest in this product. However, I am proud of this product and its role as a versatile instant camera with automatic exposure control providing low cost photomicrography capability.
Mary McCann McCann Imaging email: mccanns-at-tiac.net Tel: 617-484-7865 Fax: 617-484-2490
Nuria Cortadellas wrote: ============================================== } I have a big problem, osmium precipitates!! } does anyone can help me? ============================================== Are you sure it is "osmium" (probably OsO2 if it is of osmium composition) and not iron oxide contamination from corrosion from the tweezer tips? Vapor staining with osmium tetroxide does corrode the tips of the "normal" antimagnetic stainless steel types. And such corrosion product can migrate to a sample being supported on a TEM grid. While the chances are greater that the problem is indeed from the osmium tetroxide, that might not always be the case.
Disclaimer: Our firm, SPI Supplies offers alternatives to the "normal" antimagnetic stainless steel tweezers for holding grids so we have a vested interest in making this point.
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 ############################ ==================================================
} } From: Nuria Cortadellas \ Internet: (nuriac-at-giga.sct.ub.es) } To: MICROSCOPY BB \ Internet: } (microscopy-at-sparc5.microscopy.com) } } Subject: Osmium precipitates } } ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } I have a big problem, osmium precipitates!! } does anyone can help me? } Thanks in advance } Nuria Cortadellas } Department of Electron Microscopy } University of Barcelona } } }
-------------------------------------------------------------- Peling Fong Melville Senior Scientific Assistant Interdepartmental Facilities American Museum of Natural History Central Park West at 79th Street New York, NY 10024-5192 U.S.A. ****************************** E-mail: peling-at-amnh.org Work #: (212) 769-5469 FAX #: (212) 769-5495
After discussed dangers of the use of DMSO suggested by Wil Bigelow, I have to share with you the old knowledge that the same effect on small burns can be obtained by a simple use of the egg white. You remove the membrane lining the egg shell and put it on the burned area. If needed, you add more of these membranes. It stops pain immediately and prevents blister formation. The method is apparently known for centuries and around the world, as the egg white treatment was also described by Gabriel Garcia Marquez in "One Hundred Years of Solitude". I never experienced, nor heard about, any negative effect.
I have an older TEM with many extras for which a new home is needed. This is a Philips EM301 with both the high resolution stage and the goniometer stage. It is presently owned by a friend whos husband died before he could finish putting his lab together. The price is negotiable and the instrument, like most of the older Philips microscopes is quite servicable. If you have any interest or questions, please contact me via e-mail.
Thank you. ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^ Alexander Greene Scientific Instrumentation Services, Inc. Number 499, Post Office Box 19400 Austin, Texas 78760 Phone: 512/282-5507 FAX 512/280-0702
REASONABLY PRICED ELECTRON MICROSCOPE REPAIR ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
I would like to add to Linda's recommendations that this precipitate phenomenon may be most common in EM but it is less obvious than lead precipitates. It was published (don't ask where) over ten years ago. It's a triangle which requires phosphate, GA and Os. If no free GA remains after thorough rinsing with buffer, the Os will not result in the precipitate. A rinse which would remove precipitate (prior to the sections irradiation by an electron beam) was also published, perhaps somebody else can post that, I do not remembers the detail. In the end cacodylate solved the precipitation problem and rinsing between GA and Os is not required. It also causes no precipitation with Ca (seawater) and results in better preservation. Pity is, because phosphate buffer is o.k. to drink, whereas cacodylate is an arsenic compound which is toxic and is a carcinogen. I believe reasonable facilities and good working habits can make it quite safe to use. 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
---------- } Reply to: RE} osmium precipitates } } Dear Nuria, } Sounds like your phosphate buffer interacting with the osmium may be the cause } of precipitates. We generally fix for resin embedding in cacodylate buffer. If } phosphate buffer is used for the fixative, we wash it away 3 x 10 minutes with } cacodylate buffer then proceed with osmication. The osmium is also never } diluted in phosphate buffer. We keep a stock solution of 4% made in water then } dilute to 2% with cacodylate buffer when ready to use. Hope this helps. } Linda Chicoine } Center for Cell Imaging } Dept. of Cell Biology } Yale University
Hi Jolanta,
My recipe is (and it is the recipe from Polysciences who sells all the LWs : For K4M it is : Crosslinker A : 3.6 ml ( or 2.7g) Monomer B : 25 ml ( or 17.3 g) Initiator C : 100mg (and NOT 20.1g)
They say : 1 weigh out, into a tared vial, the crosslinker and the monomer. Mix gently by one of the following methods for three to five minutes : =
-bubble a continuous stream of dry nitrogen gas into the mixture with a Pasteur pipette. The nitrogen stream will mix the resin, and at the same time it will prevent the incorporation of oxygen. -mix gently with a glass rod. -if the vial has a small cap or lid, slowly rock the covered vial from side to side, avoiding the formation of air bubbles or foaming. Add the initiator and continue mixing until the initiator is completely dissolved in the resin. The mixture is given for ultraviolet polymerization from -50=B0C to 0=B0C= =2E Above 0=B0C, the initiator C should be replaced by the same amount of benzoin ethylether.
IN ALL THE LWs MIXTURES, YOU ONLY USE 100 TO 150 MG OF INITIATOR
You might wish to ask EMS for their booklet on the use of lowicryl and their lowicryl letters. =
My recipe is (and it is the recipe from Polysciences who sells all the LWs : For K4M it is : Crosslinker A : 3.6 ml ( or 2.7g) Monomer B : 25 ml ( or 17.3 g) Initiator C : 100mg (and NOT 20.1g)
They say : 1 weigh out, into a tared vial, the crosslinker and the monomer. Mix gently by one of the following methods for three to five minutes : =
-bubble a continuous stream of dry nitrogen gas into the mixture with a Pasteur pipette. The nitrogen stream will mix the resin, and at the same time it will prevent the incorporation of oxygen. -mix gently with a glass rod. -if the vial has a small cap or lid, slowly rock the covered vial from side to side, avoiding the formation of air bubbles or foaming. Add the initiator and continue mixing until the initiator is completely dissolved in the resin. The mixture is given for ultraviolet polymerization from -50=B0C to 0=B0C= =2E Above 0=B0C, the initiator C should be replaced by the same amount of benzoin ethylether.
IN ALL THE LWs MIXTURES, YOU ONLY USE 100 TO 150 MG OF INITIATOR
You might wish to ask EMS for their booklet on the use of lowicryl and their lowicryl letters. =
} I desperately need to convert "old spectra files" from } old systems to the newer one Intel/Windows platform and from/to Edax {--} Link !
Dear Andrea, Here at Mektech we manufacture MS Windows based EDS system that connects to Link AN10000 pulse processor. It can also read Link AN10000 and eXL spectra. For more info visit our website at www.visionol.net/~mektech or contact as directly.
I have been asked to study electrodeposited copper microstructure. My goal is to determine the relationship among electrodeposition process operational variables (electrolyte concentration, impurities deposition, aditives, current intensity applied etc.), microstructure (texture, grain size...), and mechanical properties of the metal.
You are all welcome to visit 5 copper micrographs I have posted on the Internet and please fell free to make any comments on them:=20
http://metallography.com/marti.htm
Any ideas will be welcome. Any reference info on the specific subject of electrodeposited copper microstructure will also help me. Can you tell me where to find copper cathode micrographs?
Please reply to Juan Marti at: jmartip-at-cepade.es
The descriptuion of the pictures is as follow:
The 5 micrographs correspond to the same cathode sample and show different structures among which I am not able to determine which one is the real one. I hope that you may help me in the interpretation of these images:
- Bottom left picture (cobre4). Etched with HNO3 (25%) at 70=BAC during only 5 seconds. Longitudinal section of the cathode showing what seems to be the grain boundaries. Lens Objective: 50x.
- Upper left and right pictures (cobre1). Etched with HNO3 (25%) at 70=BAC during 15 seconds. Longitudinal section of the cathode. It apparently shows big irregular grains but I=92m not sure if the sample might be under etched, thus not showing the real microstructure. Lens Objective: 50x.
- Middle right picture (cobre3). Etched with HNO3 (25%) at 70=BAC. More etching time. Longitudinal section of the cathode showing much smaller grains. Grains also seem irregular. Lens Objective: 50x.
- Middle left picture (cobre2). Etched with HNO3 (25%) at 70=BAC. Transversa= l section of the cathode showing what appears to be large grains grown in the current flow direction. Lens Objective: 20x.
- Bottom right picture (cobre6). Shows detail of abnormal grain size. Lens Objective: 20x.
Among the first 3 micrographs (cobre4, cobre1 and cobre3) can you tell which one is the real pure copper microstructure?
Any ideas will be welcome. I would also appreciate any help on specific references to copper cathode microstructures descriptions and micrographs.= =20
Thanks in advance.
Please reply to Juan Marti at: jmartip-at-cepade.es
Micrographs web site at: http://metallography.com/marti.htm
Biological Physics. Postdoctoral position at the Department of Molecular Physiology and Biological Physics of the University of Virginia School of Medicine. The research projects in which the applicant is expected to play a major role involves electron energy loss spectroscopy and energy filtered scanning transmission electron microscopy. The position will include both development of software and instrumentation for achieving 2 to 3 nm spatial resolution compositional imaging and hands-on application of the method to significant biological problems. The laboratory has been engaged in NIH- supported research developing and applying analytical electron microscopy for 25 years through an interdisciplinary program based on the collaboration between physicists and biologists. Equipment available includes a 200kV electron microscope equipped with field emission gun (Philips CM200-FEG), a GATAN electron spectrometer adapted to our own CCD camera, a CM12 electron microscope and two energy dispersive X-ray detectors. Investigators in the program are also members of the Center for Structural Biology of the University of Virginia and have programs involving collaborations with the X-ray crystallography and atomic force microscopy groups and with molecular biologists and investigators engaged in other biological disciplines.
Candidates should have a Ph.D. in Physics, material science or engineering and be familiar with computer programming, instrumentation and, preferably, experience with electron microscopy and electron energy loss spectroscopy. Applications with biographical sketch, bibliography and the names of three references should be sent to: Dr. Andrew P. Somlyo, Department of Molecular Physiology and Biological Physics, University of Virginia, P.O. Box 10011, Charlottesville, VA 22906-0011, USA. The University of Virginia is an Equal Opportunity/Affirmative Action Employer.
I apologize for the false start. It's for the Philips EM201/300/301 series microscopes we need (NOT the 200 or 400, as I thought earlier). Again, used, simple, single tilt, is fine, if not bent. Key parameter is inexpensive.
DGCollins-at-lbl.gov (510) 486-7859, or DCollins 2841 Kinney Dr, Walnut Creek, CA 94595 (510)939-2006
There is a program (NTRANS) in the MSA Software library which runs on your PDP 11 computer and can do SOME of the work you request. The software is free, but you will have to get someone to compile it. Alternatively, you ask the manufacturers to supply to you a copy of their program which translates their data into the MSA/MAS Standard Spectral File Format. This should be even more effective, as each manufacturer only need to write a R/W subroutine for their format to/from the one standard. The advantage here is that the manufactures may already have the translator compiled and running on each of the platforms that you already have. The other option is DTSA which is a National Institute of Standards and Technology (NIST) computer program for XEDS , which has many translators built in. That program however must be purchased from NIST. (Disclaimer: I have no financial interests in DTSA).
If you want a Copy of NTRANS you can get it here;
The anonymous ftp server address is
Host: ftp.msa.microscopy.com UserId: anonymous Passwd: your email address
or you can go to the master ftp site
Host: ftp.amc.anl.gov UserId: anonymous Passwd: your email address
Go to the public directory, find the MMSLib Go to the XEDS directory Go to the NTRANS directory
All the files are in there...
Here is a copy of the on-line abstract file
Ntrans.abs.
Title :NTRANS Keywords :XEDS, EELS Computer :DEC VAX 11/730-785, DEC PDP 11/2-11/73 Operating System :VAXVMS, RT-11 Programming Language :Fortran IV Hardware Requirements :None Author(s) :Nestor J. Zaluzec Correspondence Address :Argonne Nat. Lab, Electron Microscopy Center,Bldg 212 :Materials Science Division, Argonne, Illinois 60439, Abstract:
NTRANS is a computer program which translates manufacturers XEDS and EELS spectral data into the EMMPDL data format. It utilies the RWEMMPDL subroutines contained in the EMMPDL. At present this version translates both EDAX and TRACOR-Northern and Link Systems data files. Examples of translated spectra can be found spectra can be found in the XEDS and EELS subdirectories of the EMMPDL. -------------------------------------------------------------------------------
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} I buffered the osmium solution with phosphate and cacodylate } buffer and the osmium precipitates appear in the sample
Residual gluaraldehyde is famous for causing Os04 precipitates. Although you say that you rinse 4-5x, try rinsing more to be sure that you are removing all GA. Also, if your specimen size is too large, the GA may not be diffusing out of the tissue entirely because of the great distance.
(Just my two cents worth),
Good luck. ______________________________________________________________________ Donald L. Lovett e-mail: lovett-at-tcnj.edu Assoc. Professor, Dept. of Biology voice: (609) 771-2876 The College of New Jersey fax: (609) 771-2674 Trenton, NJ 08650-4700
Having spent several years wordering just how many different formats Link could possibly come up with for storing X-ray spectra, I was just a little bit enraged when they changed things AGAIN with the introduction of the ISIS system. The converters Nestor has pointed out won't work with that system.
I'd like to echo Nestor's suggestion to the manufacturers about them supplying converters and it seems that this forum could be a useful place to gather a weight of opinion. I know that there are many parameters that are sometimes stored along with the 'raw' data which would make the free translation of data formats somewhat perilous or difficult, but it ought to be possible for access to most of the data to be a darned sight easier than it is now.
Dr Simon Dumbill AEA Technology Tel: +44 1235 434245 220, Harwell Fax: +44 1235 435941 Didcot Email: Simon.Dumbill-at-aeat.co.uk Oxfordshire OX11 0RA UK
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Andre'
There is a program (NTRANS) in the MSA Software library which runs on your PDP 11 computer and can do SOME of the work you request. The software is free, but you will have to get someone to compile it. Alternatively, you ask the manufacturers to supply to you a copy of their program which translates their data into the MSA/MAS Standard Spectral File Format. This should be even more effective, as each manufacturer only need to write a R/W subroutine for their format to/from the one standard. The advantage here is that the manufactures may already have the translator compiled and running on each of the platforms that you already have. The other option is DTSA which is a National Institute of Standards and Technology (NIST) computer program for XEDS , which has many translators built in. That program however must be purchased from NIST. (Disclaimer: I have no financial interests in DTSA).
If you want a Copy of NTRANS you can get it here;
The anonymous ftp server address is
Host: ftp.msa.microscopy.com UserId: anonymous Passwd: your email address
or you can go to the master ftp site
Host: ftp.amc.anl.gov UserId: anonymous Passwd: your email address
Go to the public directory, find the MMSLib Go to the XEDS directory Go to the NTRANS directory
All the files are in there...
Here is a copy of the on-line abstract file
Ntrans.abs.
Title :NTRANS Keywords :XEDS, EELS Computer :DEC VAX 11/730-785, DEC PDP 11/2-11/73 Operating System :VAXVMS, RT-11 Programming Language :Fortran IV Hardware Requirements :None Author(s) :Nestor J. Zaluzec Correspondence Address :Argonne Nat. Lab, Electron Microscopy Center,Bldg 212 :Materials Science Division, Argonne, Illinois 60439, Abstract:
NTRANS is a computer program which translates manufacturers XEDS and EELS spectral data into the EMMPDL data format. It utilies the RWEMMPDL subroutines contained in the EMMPDL. At present this version translates both EDAX and TRACOR-Northern and Link Systems data files. Examples of translated spectra can be found spectra can be found in the XEDS and EELS subdirectories of the EMMPDL. -------------------------------------------------------------------------------
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After read all the posted messages about Glutaraldehyde and Formaldehyde with their hazadous fumes, we Electron Microscopy Sciences want to remind you all that we have been introducing in our catalog, in the Safety Section, the LAB-AIR System - an Electronic Air Purifiers, which complied with OSHA Regulations and minimizes Occupation Exposure To Toxic Vapors. The System that destroys the odors and fumes, and doesn't just mask them. Electronic Air Purifiers produce a controlled level of Ozone (O3) electrically by converting molecules of Oxygen (O2) into molecules of Ozone (O3).. Ozone, sometimes called activated oxygen, as part of the process of returning to oxygen, casts off its extra atom. That extra atom combines with the molecule of the odor's source and thereby destroys the odor by oxidation. Once Ozone's extra atom is consumed fresh air is leftbehind which was created by a natural process. For instance: HCHO + O3 = HCOOH + O2 Formaldehyde Ozone Formic acid Oxygen HCOOH + O3 = CO2 + H2O + O2 Formic acid Ozone Carbon dioxide * Water* Oxygen* * All Harmless Gases. We are not intended to introduce our product on the site, but we thought this messages are helpfull to all of our Scientists and Technicians, whose is dealing with chemicals daily in theirs enclosed labs. For more information, please contacting us at 1 800 523 5874
Electronic microscopy by transmission (Zeiss EM109)
We performed more than 20,000 photographies on not-perfored rollfilm 70 mm as AGFA Scientia, AGFA Rapidoline, AGFA Aviortho, KODAK Kodalith. Unfortunately the production of all these rollfilms are now stopped , as well as Ortho AGFA films (sheet and roll 120/135).
Is anybody has suggestions about the replacement of these high specific products ?
In fairness to Oxford, it should be mentioned that the ISIS may have its own new file format for spectra, but it does read and write the earlier eX-L and AN10000 formats, as well as the EMSA/MAS format.
The x-ray analyser manuals for the AN10000 and the eX-L both contain detailed descriptions of the spectrum file formats in appendices. The MSDOS convert program, available on eX-L's and later AN10000's (those with 3.5" floppy's, I think) will copy the spectra to MS-DOS disks, from which a simple program will readily convert them to text files.
I have written such a program. It is available by anonymous FTP from IMAGES.MIT.EDU. There are several files there, but the readme files explain what is what. In addition to the conversion program above, there is a program to duplicate the MSDOS Convert program (by reading Genie/DEMON 3.5" disks on the PC) and a program to extract images from studies.
Hope this is useful.
Tony Garratt-Reed
**************************************************** **************************************************** ** ** ** Anthony J. Garratt-Reed ** ** Room 13-1027 ** ** Center for Materials Science and Engineering ** ** Massachusetts Institute of Technology ** ** 77 Massachusetts Avenue ** ** Cambridge, Massachsetts 02139-4307 ** ** U. S. A. ** ** ** ** Phone: 617-253-4622 ** ** Fax: 617-258-5286 or 617-258-6478 ** ** ** **************************************************** ****************************************************
Please be advised of the following position opening. Contact John May at the address below.
Bob Wise
} Date: Mon, 16 Jun 1997 10:55:14 -0500 } From: "John F. May" {shayala-at-fdldotnet.com} } Subject: Electron microscopy position } X-Sender: shayala-at-pop.fdldotnet.com } To: wise-at-vaxa.cis.uwosh.edu } Cc: jedunphy-at-mariancoll.edu } X-Mailer: Windows Eudora Light Version 1.5.4 (32) } } Dr. Wise, } } I would like to alert you to a FULL TIME faculty position in Biology } at Marian College in Fond du Lac for the 1997-98 school year. We want } someone to teach the following courses: } } Bi/PhS 331 Transmission Electron Microscopy (2 cr.) } Bi 100 Life Systems (lecture only) (3 cr.) } Bi 100 Life Systems lecture & lab (evening section) } (4 cr.) } Sci 101 Integrated Physical/Biological Science (3 cr.) } (team-taught with a Phsycal Science } faculty member) } OR } Bi 201 Anatomy & Physiology (4 cr.) } } } The Spring semester schedule would be similar, with Sci 102 and Bi 202 being } offered as the second semester of those courses. The faculty member would } be expected to teach the Biology portion of the integrated course. } } Our normal course load is 24 hours per year. } } Please share this announcement with anyone you feel would be interested or } colleagues who might know a potential applicant. Have interested } individuals contact me be e-mail or phone. } } Thank you. } } John F. May, Ph.D. } Biology Coordinator } Marian College } Fond du Lac, WI 54935 } Phone: 414-923-7646 } e-mail: shayala-at-fdldotnet.com } }
Ozone sounds good in theory. I'd certainly prefer it to formaldehyde, but is that a "Hobson's choice" (choosing between the lesser of two evils)? Can you comment on the effects of ozone on lung tissue?
} To all, } } After read all the posted messages about Glutaraldehyde and Formaldehyde with } their hazadous fumes, we Electron Microscopy Sciences want to remind you all } that we have been introducing in our catalog, in the Safety Section, the } LAB-AIR System - an Electronic Air Purifiers, which complied with OSHA } Regulations and minimizes Occupation Exposure To Toxic Vapors. The System } that destroys the odors and fumes, and doesn't just mask them. } Electronic Air Purifiers produce a controlled level of Ozone (O3) } electrically by converting molecules of Oxygen (O2) into molecules of Ozone } (O3).. Ozone, sometimes called activated oxygen, as part of the process of } returning to oxygen, casts off its extra atom. That extra atom combines with } the molecule of the odor's source and thereby destroys the odor by oxidation. } Once Ozone's extra atom is consumed fresh air is leftbehind which was created } by a natural process. } For instance: } HCHO + O3 = HCOOH + O2 } Formaldehyde Ozone Formic acid Oxygen } HCOOH + O3 = CO2 + H2O + O2 } Formic acid Ozone Carbon dioxide * Water* Oxygen* } * All Harmless Gases. } We are not intended to introduce our product on the site, but we thought this } messages are helpfull to all of our Scientists and Technicians, whose is } dealing with chemicals daily in theirs enclosed labs. } For more information, please contacting us at 1 800 523 5874 } } Bang Nguyen } Electron Microscopy Sciences } } Dr. M. Dennis Goode Phone (301) 405-6917 Department of Zoology Fax (301) 314-9358 University of Maryland e-mail goode-at-zool.umd.edu College Park MD 20742 ************************************************************* "If the Lord Almighty had consulted me before embarking upon the creation, I should have recommended something simpler." -Alphonso X of Castile, 15th Century
I am currently trying to get Link to provide a simple X-Y output (channel- counts) of their spectra in tab-separated variable format from their ISIS system. That way I could easily import the spectra into Kaleidagraph or Excel. I don't care about the headers or other information; I can get that from the original data. They are "working on it", but I haven't heard anything from them for a month or so.
I'm not a programmer, but is it that hard to make an output like that? Since the spectra are plotted within their program it seems that those raw data in X-Y format MUST exist somewhere.
If they come up with something, I'll let people know.
Cheers,
John Vetrano Pacific Northwest National Laboratory _______________________________________________________________________________
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Having spent several years wordering just how many different formats Link could possibly come up with for storing X-ray spectra, I was just a little bit enraged when they changed things AGAIN with the introduction of the ISIS system. The converters Nestor has pointed out won't work with that system.
I'd like to echo Nestor's suggestion to the manufacturers about them supplying converters and it seems that this forum could be a useful place to gather a weight of opinion. I know that there are many parameters that are sometimes stored along with the 'raw' data which would make the free translation of data formats somewhat perilous or difficult, but it ought to be possible for access to most of the data to be a darned sight easier than it is now.
Dr Simon Dumbill AEA Technology Tel: +44 1235 434245 220, Harwell Fax: +44 1235 435941 Didcot Email: Simon.Dumbill-at-aeat.co.uk Oxfordshire OX11 0RA UK
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Message-Id: {1.5.4.32.19970616180604.0068f0ac-at-pop.fast.net} X-Sender: goldmrkr-at-pop.fast.net (Unverified) X-Mailer: Windows Eudora Light Version 1.5.4 (32) Mime-Version: 1.0 Content-Type: text/plain; charset="us-ascii"
Colleagues -
A friend of mine is interested to sell his microscope, so I said I would put it on the server since he is not a member. Please contact me if you have an interest and I'll pass the word.
Thanks and regards, Don Cox
---------------------------------
CARL ZEISS RESEARCH MICROSCOPE (No model # that I can find---but it is their standard research microscope body that they carried for years. Probably bought in the mid 70's. } } Trinocular head with beam splitter. } } Condenser (phase and darkfield) } } Seven (7) zeiss lenses: } 16X /0.40 Neofluar, (phase) } } 40X /0.65 Plan } } 40X /0.65 Plan (phase) } } 40X /1.0 Oil (APO) Iris Diap. (0.6-1.0) } } 100X /1.25 Oil Iris Diap. (1.25-0.8)
Hello, We are starting a histochemical EM study on acetylcholinesterase activity in brain tissue. We are searching for a method of quantifying the levels of AChE activity. I would appreciate any information from an experienced researcher, and any appropriate references.
Recently, I am having problems when sectioning my blocks. I work on=20 animal tissue, a sea pansy which is a coelenterate. I only use the polype.
MY PROBLEM: when I section, I only get the resin and where my tissu should= =20 be I have a hole!! I never had this problem. Could it be a deshydratation too long (10 min. for each ethanol, 2*10 for the 100% and= =20 2*15 for oxyde propylene.. Or residual oxide propylene..Or could it be the= =20 inclusion in agarose. My resin is hard but when I come close to the tissue it is smooth.
Is it possible to recuperate these sections? and how
My PROTOCOLE BRIEFLY: I fix my tissue in glutaraldheyde or=20 paraformaldheyde 4% (various=20 fixation I use. ) After fixing for 3 or 4 hours, I embed the tissue in an= =20 agarose solution for 24 hours and I then section those agarose blocks=20 with a vibratome. Those sections of a thickness varying between 50 to=20 200 micrometers are incubated in 1% OsO4 for 2 hours. I then dehydrate in= =20 ethanol and embed in epon and araldite for 24 hours at 60 degrees. =09=09 =20 I hope that someone can help=20 thank you.
Natacha Benrimoh Universt=E9 de Montr=E9al benrimon-at-ere.Umontr=E9al.CA (514) 343-6111 poste 1052.
To sidetrack a bit, Hobson's choice actually means "no choice", there's only one recourse and that's it!!! Hope you don't mind a bit of ribbing, Dennis! :-) As for effects of ozone on lung tissue, I'm no chemist but as I understand it, it's a free radical and nothing good comes out of mixing those radicals with living tissue....that's why we have all those quacks toting beta-carotenes and Vitamin C & E etc. Well, that's my penny's worth of comments! :-)
Meng
On Mon, 16 Jun 1997, Dennis Goode wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Bang, } } Ozone sounds good in theory. I'd certainly prefer it to } formaldehyde, but is that a "Hobson's choice" (choosing between the } lesser of two evils)? Can you comment on the effects of ozone on } lung tissue? } } -Dennis } } } Date: Mon, 16 Jun 1997 07:46:30 -0400 (EDT) } } From: BNguyen260-at-aol.com } } To: goode-at-zool.umd.edu, Microscopy-at-sparc5.microscopy.com, } } richard.lander-at-stonebow.otago.ac.nz, KPR-at-wpo.nerc.ac.uk } } Subject: Re: Glutaraldehyde: safe limits -Reply } } } To all, } } } } After read all the posted messages about Glutaraldehyde and Formaldehyde with } } their hazadous fumes, we Electron Microscopy Sciences want to remind you all } } that we have been introducing in our catalog, in the Safety Section, the } } LAB-AIR System - an Electronic Air Purifiers, which complied with OSHA } } Regulations and minimizes Occupation Exposure To Toxic Vapors. The System } } that destroys the odors and fumes, and doesn't just mask them. } } Electronic Air Purifiers produce a controlled level of Ozone (O3) } } electrically by converting molecules of Oxygen (O2) into molecules of Ozone } } (O3).. Ozone, sometimes called activated oxygen, as part of the process of } } returning to oxygen, casts off its extra atom. That extra atom combines with } } the molecule of the odor's source and thereby destroys the odor by oxidation. } } Once Ozone's extra atom is consumed fresh air is leftbehind which was created } } by a natural process. } } For instance: } } HCHO + O3 = HCOOH + O2 } } Formaldehyde Ozone Formic acid Oxygen } } HCOOH + O3 = CO2 + H2O + O2 } } Formic acid Ozone Carbon dioxide * Water* Oxygen* } } * All Harmless Gases. } } We are not intended to introduce our product on the site, but we thought this } } messages are helpfull to all of our Scientists and Technicians, whose is } } dealing with chemicals daily in theirs enclosed labs. } } For more information, please contacting us at 1 800 523 5874 } } } } Bang Nguyen } } Electron Microscopy Sciences } } } } } Dr. M. Dennis Goode Phone (301) 405-6917 } Department of Zoology Fax (301) 314-9358 } University of Maryland e-mail goode-at-zool.umd.edu } College Park MD 20742 } ************************************************************* } "If the Lord Almighty had consulted me before embarking upon the } creation, I should have recommended something simpler." } -Alphonso X of Castile, 15th Century }
Dear All, I keep a Aloe Vera plant on my windowsill. If I get a burn, I just snip off a leaf and squeeze out the jelly onto the burn. It forms a protective skin and cools the hurt. 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
I spoke to Agar Scientific yesterday - they are our normal supply source for film. They are trying to obtain another film for me to try, but no details as yet.
We have used unperforated since 1967 (1969 personally) on Philips microscopes. We found many years ago that if the negatives were well focused and also in the darkroom then no-one could differentiate between prints from plates (as then) and 35mm film on 20 x 25 cm paper.
Not a bad idea. Aloe Vera has been used in the Tropics by many for a number of ailments. Good Ole Folk Medicine.
Leo
On Mon, 16 Jun 1997, Mary Mager wrote:
} ------------------------------------------------------------------------ } The Microscopy ListServer -- Sponsor: The Microscopy Society of America } To Subscribe/Unsubscribe -- Send Email to ListServer-at-MSA.Microscopy.Com } -----------------------------------------------------------------------. } } Dear All, } I keep a Aloe Vera plant on my windowsill. If I get a burn, I just snip off } a leaf and squeeze out the jelly onto the burn. It forms a protective skin } and cools the hurt. } 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 } }
Hello everybody! Many thanks to all of you who have replied to my question about TEM film. Here, however, is a reply I got privately, which I am going to try out. Youall might like to try it, too, but I cannot yet vouch for any results.
* * We still run 35mm film in our EM Unit for both our Philips TEM's * (CM10 and 201c) as well as our Cambridge 250 SEM. * * However; on the TEM's we had the same problem you are now * experiencing. Changing to plates, I think is a step backwards. We use * to use Kodak FGP but we that became unavailable...we switched to AGFA * COPEX Pet 10 which we found as good. Perfect in the sense that it is * as sensitive, and not perforated. * } Does it have to loaded in the dark, or will red light do? (much more } convenient). * * A RED SAFELIGHT IS ALL WE USE *
+------------------------------------------------------------------------+ | Robert H.Olley Phone: | | J.J.Thomson Physical Laboratory {direct line +44 (0) 118 9318572 | | University of Reading {University internal extension 7867 | | Whiteknights Fax +44 (0) 118 9750203 | | Reading RG6 6AF Email: R.H.Olley-at-reading.ac.uk | | England URL: http://www.reading.ac.uk/~spsolley | +------------------------------------------------------------------------+
} On the subject of LN2, does anyone know the correct first aid treatment } for a burn from this substance? } } Normally, one would use cold water to cool a (heat) burn and prevent } further tissue damage. But that seems inappropriate somehow...Hot water? } Dear Anthony, To quote from p. 41 of the Electron Microscopy Safety Handbook, 2nd Ed.: "Cryogens cause burns similar to frostbite and should be treated by warming of the affected part to body temperature." Yours, Bill Tivol
Please accept my apologies if I'm going over a well traveled path but I can't seem to find the answer to my question in my records from the list, namely: what are some of the best cameras to hook up to a TEM so that we can digitize images? We have a JEOL 100CX fitted with a YAG crystal from Fullam, a Macintosh 8500, and NIH Image for our software program. I imagine we would also end up purchasing a frame grabber such as one of the SCION boards.
But meanwhile what about a camera? I know there are quite a few out there varying widely in price and capabilities and I guess I'm wondering if anyone could help me with some feedback? I'm anticipating being able to spend between $10K and (hope hope) $20K. Any and all information will be greatly appreciated!
In 1992, we studied corneal endothelial cell damage as a result of incidental contact during ocular surgery. Although we were not concerned with the epithelium or Bowman's layer, we found a stain that worked quite well with the endothelium; Trypan Blue at a concentration of 0.2%. If it helps, please refer to my article in the 1992 EMSA proceedings, Part II, p. 1106:
"Evaluation of the Biocompatibility of Polymer Surface Modifications with the Corneal Endothelium", R. Citron, B. Tunberg, A. Yamada.
Regards,
Bob ************************* Bob Citron Chiron Vision Claremont, CA 91711 (909)399-1311 Bob_Citron-at-cc.chiron.com *************************
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I am working with the human cornea and am exploring options for tissue preparation techniques and staining procedures for LM. Sections of 0.5-1.0 microns will be used to distinguish between the epithelium and underlying Bowman's layer. I need to find the optimal embedding medium as well as staining procedure.
EDAX Offers a conversion program for the PV9900 called PVconvert. The DX-4 system allows you an option to save spectra as a .csv file for spreadsheet use.
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I am currently trying to get Link to provide a simple X-Y output (channel- counts) of their spectra in tab-separated variable format from their ISIS system. That way I could easily import the spectra into Kaleidagraph or Excel. I don't care about the headers or other information; I can get that from the original data. They are "working on it", but I haven't heard anything from them for a month or so.
I'm not a programmer, but is it that hard to make an output like that? Since the spectra are plotted within their program it seems that those raw data in X-Y format MUST exist somewhere.
If they come up with something, I'll let people know.
Cheers,
John Vetrano Pacific Northwest National Laboratory _______________________________________________________________________________
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Having spent several years wordering just how many different formats Link could possibly come up with for storing X-ray spectra, I was just a little bit enraged when they changed things AGAIN with the introduction of the ISIS system. The converters Nestor has pointed out won't work with that system.
I'd like to echo Nestor's suggestion to the manufacturers about them supplying converters and it seems that this forum could be a useful place to gather a weight of opinion. I know that there are many parameters that are sometimes stored along with the 'raw' data which would make the free translation of data formats somewhat perilous or difficult, but it ought to be possible for access to most of the data to be a darned sight easier than it is now.
Dr Simon Dumbill AEA Technology Tel: +44 1235 434245 220, Harwell Fax: +44 1235 435941 Didcot Email: Simon.Dumbill-at-aeat.co.uk Oxfordshire OX11 0RA UK
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What are the people at Marion College thinking? They want someone to teach electron microscopy and three other courses in one semester? I have taught a 4-hour course in electron microscopy for 10 years. The first 7 I was on my own and was spending better than 50 hours a week just with that course. The last three years I have had an assistant and the two of us stay very busy. If electron microscopy is taught hands-on, it is a labor intensive course for both the students and the teachers, and the only way it can be a useful and meaningful course is to be hands-on. That is what it is all about. One can read all the theory in the world, but nothing substitutes for sitting in front of an ultramicrotome for a few hours a day and putting both hands on all those neat controls on the electron microscope.
You are nearly certainly experiencing problems with tissue falling out of the section because of suboptimal infiltration. You probably have a very hard item which is difficult to infiltrate. Epon-Araldite combinations are very viscous, but the adhesive qualities of Araldite make it a good choice in these cases. Try infiltration with propylene oxide and resin 2:1 for one hour, 1:1 for 2 hours, 1:3 for 3 hours. Then pure resin for one hour. Then pure resin (new tube) for overnight. In the AM change resin again and rotate for another 2 hours. Your specimen vials must be in motion the entire time that infiltration is taking place. If your problem is not solved this way, please call or E-mail me. There may be other influences at work here. I assume that you have a sharp diamond knife at the correct angle, etc. Bye, Hildy Crowley hcrowley-at-DU.edu
I agree 200% with you on this Joyce. It's about time some of these administrators are set straight regarding EM. True it is a tool, but a very labor intensive one both to learn and to use. I just rubs me the wrong way to hear of all the downsizing and putting undertrained, undereducated people in charge of EM labs. I can't help but howl when I see an EM position requiring extensive knowledge and training in many sophisticated techniques for a $22,000 annual salary.
my $00.02 worth ed
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 } } What are the people at Marion College thinking? They want someone to } teach electron microscopy and three other courses in one semester? } I have taught a 4-hour course in electron microscopy for 10 years. The } first 7 I was on my own and was spending better than 50 hours a week } just with that course. The last three years I have had an assistant and } the two of us stay very busy. } If electron microscopy is taught hands-on, it is a labor intensive } course for both the students and the teachers, and the only way it can } be a useful and meaningful course is to be hands-on. That is what it is } all about. One can read all the theory in the world, but nothing } substitutes for sitting in front of an ultramicrotome for a few hours a } day and putting both hands on all those neat controls on the electron } microscope.
Some months ago I asked for suggestions regarding the best way to handle mechanical pump oil vapors. We had a problem with pumps working in a room with people and computers. I received many helpful replies and thought I should share our experience.
First, it is the unanimous opinion of everyone who responded and with all those I checked with that exhausts from mechanical pumps and people do not mix. For reasons of health, fire safety, cleanliness and liability, pump exhaust should be controlled.
Second, many filters designed to stop oil mist from pumps may not be sufficient to capture all the vapors and/or they may need more frequent changing to be effective, ie if you smell oil vapor, filter or not, you need to fix it.
Third, the best solution to the problem is to vent the pumps to the outside or at least to the exhaust ventilation of the building.
So, here is how we tried to solve our problem of controlling pump exhaust. I proposed that the pump exhausts be connected to the building ventilation system. I did not get too far with this plan. I got a lot of resistance, mostly based on the expense involved with ducting (it is about 50' to the nearest fume hood) and rebalancing the air circulation system of the entire 5 floor building. The story was that it would take an incredibly long time to get anything like that done because it would involve getting architects and engineers to plan and spec the project etc.
I tried to enlist the help of our health& safety office but they couldn't do much because there seem to be no guidelines to use to determine if we were violating health standards. It was sort of a Catch-22, they agreed oil mist was probably not healthy, but they could not use their leverage to demand building modifications because there were no standards to enforce.
So, it was back to the drawing board for a solution. Several replies suggested using PVC pipe to make a pathway for the vapors either to the outside or to the nearest fume hood. I was already to try that when the campus fire dept. vetoed the use of PVC pipe for any kind of pump exhaust.
By now I had made good friends with one of the campus plumbers who was trying to help me with the job. He found an acceptable flame resistant hose and a large wall mounted filter that should do the job. The filter is supposed to remove all oil vapors and it has a pressure gauge to indicate when the filter should be changed. Only time will tell if this is a good solution to our problem, so far, so good.
Correcting the problem of mechanical pumps discharging oil mist into the air turned out to be more of a problem than I had anticipated. We are all breathing a lot easier in our lab now and I would encourage everyone to at least check their pump exhaust situation to make sure that it is adequately controlled.
Jonathan Krupp Microscopy and Imaging Lab University of California Santa Cruz, CA 95064 (408) 459-2477 FAX (408) 429-0146 jmkrupp-at-cats.ucsc.edu
COMPUTER INTERACTIVE HIGH-RESOLUTION TRANSMISSION ELECTRON MICROSCOPY
N.C.E.M, LBNL, Berkeley, California
The National Center for Electron Microscopy announces its fourth ANNUAL SUMMER SCHOOL on COMPUTER INTERACTIVE HIGH-RESOLUTION TRANSMISSION ELECTRON MICROSCOPY, including Image Acquisition, Image Processing and Image Simulation, to be held at the National Center for Electron Microscopy during the week of August 25-29, 1997.
The aim of the School is to train participants in the techniques of computer-assisted high-resolution electron microscope image acquisition and image interpretation, including remote-control microscopy. Participants will learn general principles and apply them to specific cases. Participants will be taught the use of computers to obtain images on NCEM microscopes, followed by training in the use of application programs for image interpretation by image processing and image simulation. Participants wanting to apply school techniques to their own projects will be encouraged to extend their visit to NCEM into the next week -- note that this requires a proposal be submitted with advance notice sufficient for project approval.
For more information, please see - http://ncem.lbl.gov/NCEM/workshops.html
:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:. Michael A. O'Keefe, Deputy Head National Center for Electron Microscopy Lawrence Berkeley National Laboratory University of California Berkeley, California 94720 tel: (510) 486-4610 fax: (510) 486-5888 email: maok-at-lbl.gov http://ncem.lbl.gov/ :.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.:.
Request for Marine phytoplankton preparation protocol for scanning electron microscopy.
I am working on the structure of phytoplankton (Marine Chlorella, Isochrysis, Tetraselmis, Chaetocerus) before and after cryopreservation.
At present I am using this protocol 1. a. centrifuge the phytoplankton samples b. add ammonium formate to remove salts crystal. c. fix in 4% buffered gluteraldehyde for 12 to 24 hrs at 4 =B0C
2. a. washing with distill water for 3 changes of 10 minutes =09 each.
3. a. Fix in 1% buffered Osmium tetroxide for 2 hrs at 4 =B0C
4. a. Rinse in distill water for 3 changes of 10 minutes each.
5. Dehydration in a series of acetone a. 35% 5 minutes b. 50% 5 minutes c. 75% 5 minutes d. 95% 5 minutes e. absolute acetone 15 minutes, 3 changes
6. Viewing under electron microscope (JEOL 6400) The cell deformed/shrunken
Can you suggest other protocol for marine phytoplankton.=20
Your suggestion is very much appreciated.
Thank you.
Hishamuddin Omar Department of Biology Faculty of Science and Environmental Studies Universiti Putra Malaysia Malaysia
Hi all, I am after some advice on the care of the FE tip and vacuum on a Hitachi 4500.
1. How often should one bake out? The manual recommends baking out when the vacuum deteriorates. After about 8 months operation ours is better than it was to start with - IP1&2 off-scale, IP3 at 7x10-7 Pa. On the other hand many people seem to recommend baking at fairly short intervals "whether it needs it or not". We are inclined to a non-interventionist approach but are getting a bit nervous...any advice?
2. What should the flash current intensity be? Ours started at around 15-20 (and we sometimes flashed twice to get a reading in the high twenties) but has steadily crept up and is now in the high forties. Is this good, bad or indifferent? Is it perhaps related to question 1? If it gets too high does it wreck the tip? We are generally flashing once or twice a day.
cheers 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
It seems like you should't worry so much...the Hitachi FE-SEMs typically run many years under your conditions before tip replacement. Our experience is with a 10-year old S-800 and a 5-year old S-4500. The first emitter on the S-800 lasted 62 months, and the second is still working perfectly 61 months later. The first S-4500 emitter is, interestingly, 61 months old at present, and it still has the same flash characteristics as it started with...the flash current is in the mid-forty range. It is typically flashed once a day, or perhaps twice if the operation extends into the evening. I am not aware of any bake-outs of the gun except for the extensive (~10 day?)initial bake-out upon installation. Anecdotally, I have heard of Hitachi FE-SEMs whose emitters lasted in the 8 year range...perhaps you will have responses from operators of some of those instruments also with their experiences.
Larry
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Dr. Lawrence F. Allard Senior Research Staff Member High Temperature Materials Laboratory Oak Ridge National Laboratory 1 Bethel Valley Road Bldg. 4515, MS 6064 PO Box 2008 Oak Ridge, TN 37831-6064
My E-Mail system has crashed a couple of times lately, and I believe that I have lost some messages. First, based on OHSS (Occupational Health and Safety Standards) provides that no employee should be in an environment where the ozone level on average is more than 0.1 parts-per-million for more than 40 hours per week or more than average of 0.3 parts-per-million for more than 15 minutes at any one time. ( Because of the limitation of the site, we can not display a chart , which is shows the Human Tolerance for Ozone). The LAB AIR Units are designed to produce the ozone level less than the limitation, and to operate within the OHSS guidelines. You should know that even with strong odor, the amount of ozone required to mask them out is only approximetely 0.04ppm, medium odor approx. 0.03ppm and light odor approx. 0.02ppm. Secondly, You are not breathing ozone air, you're breathing normal air, the Lab-Air turns on only when needed and ozone air is produced by the lab air just enough to mask (oxidizing) the odor sources. For instance, you're drinking water, not drinking chlorine, but in the water that you are drinking has some amount of chlorine, which is used to remove bacterias. When you do an embedding mixture, for instance !00ml Araldite-Epon mixture, you are using only maximum 1.5% of DMP-30 (1.5ml) to make the whole 100ml ot that mixture turn into a solid plastic block. Back to ozone air, you need just a small amount of O3 to oxidize the unwanted odor or unwanted chemicals which presence inside the room. Each Lab-Air has the timer to set the length of time which you want the Lab-Air to work, as well as setting for ozone to control the ozone air output, but the ozone output never exceeds the limit which is the Human Tolerance for Ozone. Thirdly, the O3 is unstable, which means it has a very short life, by its very nature Ozone will revert to oxygen within a short period of time.