would like to subscribe to the microscopy listserver
kind regards,
Michael Bosma ****************************************************************************** * * * Michael Bosma Telephone: +46 418 670 62 * * The Swedish University of Fax: + 46 418 670 81 * * Agricultural Sciences E-mail: Michael Bosma-at-vf.slu.se * * Dept of Plant Breeding Research * * S-26831 Svaloev, Sweden * * * ******************************************************************************
For the record, over a period of time the various BioRad consumables ended up first at Energy Beam Sciences, then at SPI Supplies. Please contact both of these companies with any questions about products from the old BioRad catalog. Steven Slap, Vice-President Energy Beam Sciences, Inc.
Dear colleagues, can someone help by providing a good recipe for reducing osmium tetroxide to help increase contrast in epoxy resin - embedded biological material? Is potassium ferri-, or ferro- cyanide the best? I thought I knew the answer but a post-doc here has given me doubts so we have a small wager riding on this.
Message-ID: {edickg.1165621758C-at-mail.its.rpi.edu} Gary Login {glogin-at-bih.harvard.edu}
John Mansfield responded to Gary Logins' post by asking, "Although not from a company this looks much like a shameless commercial to sell this guy's books, what do you think?" Gary Login is recognized as one of the leading experts in the world on this subject. I am absolutely convinced that his references to his own articles and books were made in a sincere effort to be helpful to a fellow microscopist looking for source material on a technique. This is far from the first time a microscopist has mentioned his own publications in a posting to this list (I recall even seeing ISBN numbers). By the way, I have no commercial interest in Gary's book, which is sold by some of my esteemed competitors. Steven Slap, Vice-President Energy Beam Sciences
I've used potassium ferrOcyanide-but mainly as a membrane enhancer for neuronal tissues. If I need to just reduce overall darkening by the osmium I use 7% sucrose in the osmication-seems to slow down that process a bit. I did find that the ferrocyanide-osmium will destroy delicate tissues not well fixed in GLA-I measured the pH and as I recall it was quite high-around 10-11. You also have to be careful of the buffer system you are in-buffer incompatabilities will give you a fine crystalline precipitate which cannot be removed. Phosphate is out, cacodylate is OK. I have not tried this on any HRP-filled material-I have no idea whether or not it could damage the chromogen... Grace Kennedy..
While we're on this topic, could I ask a related question? Why does the usual ferri-ferrocyanide method (Karnovsky 1971, ASCB abstracts; or Russell and Burguet, 1978, Tissue and Cell 9:751) stain membranes so well, but leaves ribosomes virtually invisible?
Kent A. Kent Christensen, University of Michigan, {akc-at-umich.edu}
----------------------------------------
On 1 Nov 1995, Paul Webster wrote:
} Dear colleagues, } can someone help by providing a good recipe for reducing osmium tetroxide to } help increase contrast in epoxy resin - embedded biological material? Is } potassium ferri-, or ferro- cyanide the best? I thought I knew the answer but a } post-doc here has given me doubts so we have a small wager riding on this. } }
Thanks are due to Drs. Steven Slap and Gary Login who have responded to my posting . It was unfortunate that someone thought that Dr. Login was try to sell his book. I do not see that he has that intention. Dr. Login not only answered my questions but also suggested a number of reference to read. His effort must be highly commended. I am sure that the references and the answers will be useful for my friend and for anyone who wants to investigate more about this technique.
Well John, I don't think it matters much. Do you think that there is such a huge bibliography on the subject that he has purposefully refrained from mentioning other books so that he can cream off the huge profits that a greater market share would give? I don't, he seems to have answered the original question fairly.
John F. Mansfield wrote:
} Although not from a company this looks much like a shameless commercial to } seel this guy's books, what do you think? } } } 1) Rapid (i.e.,in seconds) microwave fixation is used with success to fix } } plant } } and animal tissues. In addition, several reports show the benefits of using } } microwave fixation to preserve soft tissues encased by hard shells (e.g., } } clams, } } teeth, insects). } } } } Recommended reading: } } Login, G. R., & Dvorak, A. M. (1994). Application of microwave fixation } } techniques in pathology to neuroscience studies: A review. J Neurosci } } Methods, 55, 173-182. } } Login, G. R., & Dvorak, A. M. (1994). Methods of microwave fixation for } } microscopy. A review of research and clinical applications: 1970-1992. Prog } } Histochem Cytochem, 27/4, 1-127. } } Kok, L. P., & Boon, M. E. (1992). Microwave Cookbook for Microscopists. (3rd } } ed.). Leyden: Coulomb Press. } SNIP
A colleague at our institution needs to purchase a stereomicroscope (aka "dissecting" stereo scope). Needs: magnification up to 60-80x, trinoc with Polaroid camera capable of generating positive/negatives. Price limitation is $3,000. Please contact me with information.
############################################################################# John J. Bozzola, Director Center for Electron Microscopy 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 #############################################################################
Both work, but the "ic" works stronger ( too strong sometimes , it gives uneven and "funny" nuclei if over stained.).
I Fix first in OsO4, and at the end of the incubation, I will add 3% KCN ( "ic " variety) for 10-15 minutes. Helps greatly, and the shorter- later times keep the funny nucleus effect from happening.
Lou Ann
} Dear colleagues, } can someone help by providing a good recipe for reducing osmium tetroxide to } help increase contrast in epoxy resin - embedded biological material? Is } potassium ferri-, or ferro- cyanide the best? I thought I knew the answer } but a } post-doc here has given me doubts so we have a small wager riding on this.
*********************** Lou Ann Miller Microscopic Imaging Laboratory College of Veterinary Medicine University of Illinois 2001 S Lincoln Ave Urbana, Illinois 61801 217-244-1566 lmiller-at-ux1.cso.uiuc.edu
OK, so Gary Login's post was not commercially based. I'm sorry. I was going to send the question to Nestor Zaluzec only but I got distracted and sent it to the list by mistake. However, I think it does bear some further comment and/or discussion. This list is getting very large and there are now many cases of messages that should not be sent to all subscribers without there being some indication that a large number of them want to see the responses. It really shouldnt have been posted to the list at all, and my reasoning for that is below.
Net etiquette (or Netiquette as it is typically known ) says that if there is a question in a mailing list or newsgroup then the correct response is to send a message to the original poster. Often a poster will say "private replies please, if there is enough interest I will summarize to the net/list". This should be implicit in ALL posts. In this manner if the poster receives many replies, and there is much duplication in the replies, then any summary can be edited by the original poster before sending the information out to everyone on the list/net. People wishing copies of the replies also send requests to the original poster. If the original poster geets say five to ten "me too" requests for the information You may ask why should the original poster have to do this editing and summarizing work? Well they are getting the free info courtesy of others so it isnt exactly a large price to pay.
Maybe these instructions should be part of the files that gets sent to new subscribers.
} Although not from a company this looks much like a shameless commercial to } sell this guy's books, what do you think? } } } 1) Rapid (i.e.,in seconds) microwave fixation is used with success to fix } } plant } } and animal tissues. In addition, several reports show the benefits of using } } microwave fixation to preserve soft tissues encased by hard shells (e.g., } } clams, } } teeth, insects). } } } } Recommended reading: } } Login, G. R., & Dvorak, A. M. (1994). Application of microwave fixation } } techniques in pathology to neuroscience studies: A review. J Neurosci } } Methods, 55, 173-182. } } Login, G. R., & Dvorak, A. M. (1994). Methods of microwave fixation for
John Mansfield North Campus Electron Microbeam Analysis Laboratory 413 SRB, University of Michigan 2455 Hayward, Ann Arbor MI 48109-2143 Phone: (313)936-3352 FAX (313)936-3352 jfmjfm-at-engin.umich.edu, John.F.Mansfield-at-umich.edu or jfmjfm-at-umich.edu they all reach me! URL: http://wwwpersonal.engin.umich.edu/~jfmjfm/jfmjfm.html
} John Mansfield responded to Gary Logins' post by asking, "Although not from a } company this looks much like a shameless commercial to } sell this guy's books, what do you think?" } Gary Login is recognized as one of the leading experts in the world on this } subject. I am absolutely convinced that his references to his own articles and } books were made in a sincere effort to be helpful to a fellow microscopist } looking for source material on a technique. } This is far from the first time a microscopist has mentioned his own } publications in a posting to this list (I recall even seeing ISBN numbers). } By the way, I have no commercial interest in Gary's book, which is sold by some } of my esteemed competitors. } Steven Slap, Vice-President } Energy Beam Sciences
I'm really happy about having experts in MANY fields on this list. I'll admit that I don't know much about microwave-enhanced processing, much less the folks doing it, even though I'm on the biological side of EM. I have heard recently that there have been some very significant advances in the techniques and technology, and look forward to a lot of discussions about them. I hope everyone who has useful contributions will chime in.
I stain most of my EM grids by hand but for large numbers of grids, I prefer to use the Reichert Ultrostainer. Since the Ultrostainer combines the waste into one container, I am faced with finding a disposal site that will take the combined waste. It has been difficult for us in the past to dispose of our waste, but I have just been informed that now no one will accept it. The radiation sites won't take it because of the lead and the hazardous waste sites won't take it because of the uranyl. We put absolutely nothing but water down the drain so public sewers are not an option. I will be contacting Leica to see if there is a way of separating the waste, but in lieu of that, how do the rest of you dispose of this combined waste ?? Any help will be greatly appreciated.
Barbara Hartman Schering-Plough Research Lafayette, NJ 07848
Thank you all for your replies. However, from the diverse replies, you see my problem. Here is the original reply I gave when asked: To reduce osmium from osmium (viii) to osmium (iv), the other component has be oxidised! Since iron appears in two forms Fe (iii), ferri and Fe (ii) the Fe (ii) has to be used. And Fe (ii) is ferro-. Therefore it would be reasonable to assume that ferrocyanide would be the one to use and, in fact, this is what I have been using for years with excellent results.
I am mystified why others say that the ferricyanide works just as well, and I know there are are many published papers using this from respected laboratories. This just makes it all the more mysterious, or am I missing something very simple here?
I thank everyone who replied, but special thanks must go to the contributors who supported me (without knowing it): George Ruben who said - "Potassium ferro- cyanide can reduce osmium tetroxide and in so doing becomes ferricyanide!" Ubirajara P. Rodrigues-Filho - "The reduction of osmium tetraoxide is probably by ferrocyanide. Ferrocyanide is oxidized to ferri reducing the osmium compound." And Walt Bobrowski, who provided the Karnovsky reference " Use of ferrocyanide-reduced osmium tetroxide in electron microscopy". ASCB meeting New Orleans, LA 1971:146.
Unfortunately the stakes of the wager were not so great that they could be shared.
Best regards, Paul Webster Center for Cell Imaging Yale University school of Medicine http://info.med.yale.edu/cellimg
Netiquette (I hate all those nerdy little "I'm a Net-insider and you're not" things like :), :(, BTW, and IMHO) also has clear guidelines on "flaming" (for non-nerdy Net users, this refers to complaints, particularly specific attacks on the originator of a posting). Net etiquette also says that the SysOp is king and it is therefore not your duty nor anyone else's to bash people who have not used to mailing list correctly. Lastly, experienced Net users don't make mistakes like sending replies to the list that are meant for the original user. Take your lumps (and they are roudly deserved as witnessed by the thread YOU generated) and let the SysOp decide when someone has stepped over the lines. Enough already!
} OK, so Gary Login's post was not commercially based. I'm sorry. I was } going to send the question to Nestor Zaluzec only but I got distracted and } sent it to the list by mistake. However, I think it does bear some further } comment and/or discussion. } This list is getting very large and there are now many cases of messages } that should not be sent to all subscribers without there being some } indication that a large number of them want to see the responses. It } really shouldnt have been posted to the list at all, and my reasoning for } that is below. } } Net etiquette (or Netiquette as it is typically known ) says that if there } is a question in a mailing list or newsgroup then the correct response is } to send a message to the original poster. Often a poster will say "private } replies please, if there is enough interest I will summarize to the } net/list". This should be implicit in ALL posts. In this manner if the } poster receives many replies, and there is much duplication in the replies, } then any summary can be edited by the original poster before sending the } information out to everyone on the list/net. People wishing copies of the } replies also send requests to the original poster. } If the original poster geets say five to ten "me too" requests for the } information } You may ask why should the original poster have to do this editing and } summarizing work? Well they are getting the free info courtesy of others } so it isnt exactly a large price to pay. } } Maybe these instructions should be part of the files that gets sent to new } subscribers. } } } Although not from a company this looks much like a shameless commercial to } } sell this guy's books, what do you think? } } } } } 1) Rapid (i.e.,in seconds) microwave fixation is used with success to fix } } } plant } } } and animal tissues. In addition, several reports show the benefits of using } } } microwave fixation to preserve soft tissues encased by hard shells (e.g., } } } clams, } } } teeth, insects). } } } } } } Recommended reading: } } } Login, G. R., & Dvorak, A. M. (1994). Application of microwave fixation } } } techniques in pathology to neuroscience studies: A review. J Neurosci } } } Methods, 55, 173-182. } } } Login, G. R., & Dvorak, A. M. (1994). Methods of microwave fixation for } } John Mansfield } North Campus Electron Microbeam Analysis Laboratory } 413 SRB, University of Michigan } 2455 Hayward, Ann Arbor MI 48109-2143 } Phone: (313)936-3352 FAX (313)936-3352 } jfmjfm-at-engin.umich.edu, John.F.Mansfield-at-umich.edu } or jfmjfm-at-umich.edu they all reach me! } URL: http://wwwpersonal.engin.umich.edu/~jfmjfm/jfmjfm.html
Technically not a microscopy question but an imaging question.
Does anyone out there know of any source for converting a european PAL format laserdisk (of microscopic images and audio) into any common North American format (i.e. NTSC Video tape or CD-ROM) so we can use the disk as a teaching tool? (The company that sells the disk does not offer it in NTSC or have the ability of converting it).
Greetings, John Mansfield wrote: } OK, so Gary Login's post was not commercially based. I'm sorry. I was } going to send the question to Nestor Zaluzec only but I got distracted and } sent it to the list by mistake. However, I think it does bear some further } comment and/or discussion. } This list is getting very large and there are now many cases of messages } that should not be sent to all subscribers without there being some } indication that a large number of them want to see the responses. It } really shouldnt have been posted to the list at all, and my reasoning for } that is below. } } Net etiquette (or Netiquette as it is typically known ) says that if there } is a question in a mailing list or newsgroup then the correct response is } to send a message to the original poster. ...
I disagree completely. THere is no such thing as ONE and only one proper response. Instead, there are many kinds of responses. Although some responses are likely to be really speciallized, many other responses are likely to be of interest to many more than the poster. In the case in point, its obvious from the postings that microwave fixation is of concern to many, even in passing. It's much more efficient, spontaneous and interesting for the responses of a general nature to be posted directly to the group. They all have subject lines. IF you don't want to read a post about microwave fixation-hit the delete key.
I've had a recent request for a resolution standard in the 50-100 nm range. I believe that this is for a relatively low-resolution SEMPA system, but could be mistaken. This seems to be awkwardly between the {5 nm standards used for modern SAM resolution and a good optical microscope standard. I was hoping that someone either might have an inspiring though or perhaps an idea from the days when SEM resolution wasn't quite what it is today?
Please respond directly; if anyone expresses interest I will summarize responses.
Daniel L. Callahan Department of Mechanical Engg. and Materials Science Rice University dlc-at-owlnet.rice.edu
I've had a recent request for a resolution standard in the 50-100 nm range. I believe that this is for a relatively low-resolution SEMPA system, but could be mistaken. This seems to be awkwardly between the {5 nm standards used for modern SAM resolution and a good optical microscope standard. I was hoping that someone either might have an inspiring though or perhaps an idea from the days when SEM resolution wasn't quite what it is today?
Please respond directly; if anyone expresses interest I will summarize responses.
Daniel L. Callahan Department of Mechanical Engg. and Materials Science Rice University dlc-at-owlnet.rice.edu
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At 11:34 PM 11/1/95 -0400, John F. Mansfield wrote:
} Net etiquette (or Netiquette as it is typically known says that if there } is a question in a mailing list or newsgroup then the correct response is } to send a message to the original poster. Often a poster will say "...... I will summarize to the net/list....." *************
It seems to me that your approach would hamper the free exchange that this list is so good at and would be a lot more work than merely erasing the superfluous replies; it's not that difficult. People seem to be pretty skilled at crafting their SUBJECT: headings so you don't have to read everything posted. * * Joiner Cartwright, Jr. * *
} If you } are getting fringe contrast in the TEM, then the particles are not amorphous. } Dear Scott, Are the fringes observed when an opaque material with a flat edge occludes a light source the same as those seen in the EM at the edge of a particle? If so, then the crystallinity of the particle has no bearing on whether the fringes are there. I think all that's required is an abrupt change in mass density (or opacity). Yours, Bill Tivol
} } Dear colleagues, } can someone help by providing a good recipe for reducing osmium tetroxide to } help increase contrast in epoxy resin - embedded biological material? Is } potassium ferri-, or ferro- cyanide the best? I thought I knew the answer but a } post-doc here has given me doubts so we have a small wager riding on this. } Dear Paul, To reduce osmium, one must oxidise the other reagent, so ferri (Fe2+) should be better than ferro (Fe3+). Yours, Bill Tivol
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At 08:46 AM 11/2/95 -500, Richard Edelmann wrote: } Technically not a microscopy question but an imaging question. } } Does anyone out there know of any source for converting a european } PAL format laserdisk (of microscopic images and audio) into any } common North American format (i.e. NTSC Video tape or CD-ROM) so we } can use the disk as a teaching tool? (The company that sells the } disk does not offer it in NTSC or have the ability of converting it). } } ************** Richard -
Look in your yellow pages under { {video - tape duplicating & transfer services} } . You should find one or more, depending on the size of your community, parties who can help you.
Joiner Cartwright, Jr., Ph.D. Director, Electron Microscopy tel.: (713)798-4658 Department of Pathology, Rm.286-A FAX: (713)798-3945 Baylor College of Medicine Internet: joiner-at-bcm.tmc.edu One Baylor Plaza Compuserve: 71555,1206 Houston, Texas 77030 U.S.A.
extract from faq-at-soc.cult.german I know, it's not about laser disks, but give it a try!
PAL format videotapes (as used in Germany) will not display properly using an NTSC (used in, eg, USA) based VCR and vice-versa.
There are services where video conversion from any format to any other format can be made for a fee (VHS, VHS-C and 8 mm types of cassettes.) This will allow playback of videotapes made overseas using US TV's and VCR's (PAL, SECAM -} NTSC) and vice-versa (NTSC -} PAL, SECAM, etc ...)
It is also not too expensive to get a VCR which is able to **play** NTSC and PAL tapes. Only a few VCR's are able to **record** and play VHS tapes in NTSC and PAL (e.g. Panasonic AG-W1, about DM 5000). Cheaper VCR's are able to play different formats (NTSC, PAL, SECAM).
**Do it yourself**
With these setups you can transfer from NTSC to/from PAL at reasonable cost. Don't expect studio quality though: o Akai VS R110EM is a three system unit - PAL, NTSC, SECAM , costs about US$200 mailorder (smile video, nyc). o AKAI VSX-560, *HiFi-Stereo*, tuner, features include NTSC playback on PAL TV, US$500 (mailorder from 47th St Photo) o AIWA MG360S also 3 systems, costs about US$450 (mail order, j/R music world, nyc, 1 800 221 8180) o Another VCR that is "reasonably" priced is sold by Radio-Shack. The VCR is available through special order only; and not all Radio Shack employees know that this machine even exists. If they don't, have them look in the current catalog for #16-706. The cost is US$600. (You'll need a second VCR for conversions.) [3/94]
**Commercial conversion**
o International Video Conversion 520 Harvest Lane, Raleigh, NC 27606-2217, tel (919) 233-8689
Fees: US$25 + 5 S&H, Price of a High Grade Cassette Included, 2hrs or less. Delivery: Mailed back the next day, express shipping at request. Payment: Check, Cash or Money Order mailed with tape.
o sasjrm-at-unx.sas.com does it for US$5 per hour + US$3 for the blank tape. Formats: NTSC, PAL, NPAL, MPAL, SECAM, MSECAM
o Soffel VDO 2250 Monroe St #263, Santa Clara, CA 95050, tel (408) 985 2098
US$20 per tape (up to 2h, each add. hour US$10). Tape, S&H included. Mail only, next day shipping, overnight available. Check, cash, money order. Does: NTSC (8mm, Hi8, VHS) -} PAL (VHS)
o Give your local shops a try! I found a **Camera Shop** that does PAL {-} NTCS conversions; a bit expensive, though (US$20/h). But if you need something the very next day... [1/94]
I disagree with John's view that replies should be routinely posted privately. I learn a lot by scanning the replies, often when I've not even noticed the original query. Provided that a subject field is entered, it's very easy just to delete items which one doesn't want to read.
Ritchie
Ritchie Sims phone: 61 9 3737599 ext 7713 Department of Geology fax: 61 9 3737435 University of Auckland Private Bag 92019 Auckland New Zealand
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We're interested, man. Share it. ************
At 09:52 AM 11/2/95 -0600, you wrote: } } All: } } I've had a recent request for a resolution standard in the 50-100 nm range. } I believe that this is for a relatively low-resolution SEMPA system, but } could be mistaken. This seems to be awkwardly between the {5 nm } standards used for modern SAM resolution and a good optical microscope } standard. I was hoping that someone either might have an inspiring } though or perhaps an idea from the days when SEM resolution wasn't quite } what it is today? } } Please respond directly; if anyone expresses interest I will summarize } responses. } } Daniel L. Callahan } Department of Mechanical Engg. and Materials Science } Rice University } dlc-at-owlnet.rice.edu } }
I am also interested in locating a source of mussel adhesive protein. If anyone knows of a source, please post it on the list, since I think others might be interested. Melanie Behrens
I'm with you, Baskin! Better too many choices than too few. I like reading all the replies and drawing my own conclusions filtered through my biases rather than some one else's
} Greetings, } John Mansfield wrote: } } OK, so Gary Login's post was not commercially based. I'm sorry. I was } } going to send the question to Nestor Zaluzec only but I got distracted and } } sent it to the list by mistake. However, I think it does bear some further } } comment and/or discussion. } } This list is getting very large and there are now many cases of messages } } that should not be sent to all subscribers without there being some } } indication that a large number of them want to see the responses. It } } really shouldnt have been posted to the list at all, and my reasoning for } } that is below. } } } } Net etiquette (or Netiquette as it is typically known ) says that if there } } is a question in a mailing list or newsgroup then the correct response is } } to send a message to the original poster. ... } } I disagree completely. THere is no such thing as ONE and only one } proper response. Instead, there are many kinds of responses. Although some } responses are likely to be really speciallized, many other responses are } likely to be of interest to many more than the poster. In the case in } point, its obvious from the postings that microwave fixation is of concern } to many, even in passing. It's much more efficient, spontaneous and } interesting for the responses of a general nature to be posted directly to } the group. They all have subject lines. IF you don't want to read a post } about microwave fixation-hit the delete key. } } Just my two cents, } Tobias Baskin } } } } - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - } ___ ____ ^ ____ _____ Tobias I. Baskin } / \ / / \ / \ / University of Missouri } / | / / \ / / Biological Sciences } /___ / /__ /_____\ / /__ 109 Tucker Hall } / / / \ ( / Columbia, MO 65211 USA } / / / \ \ / voice: 314-882-0173 } / /____ / \ \____/ /_____ fax: 314-882-0123 } } } -at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at- Greg Erdos Phone: 904-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax 904-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 -at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at-
It may be Netiquette to send replies only to the originator of the question but I for one would find the value of the mailing list greatly reduced if I only saw questions not answers. I value the broad range of topics discussed and in a number of cases the answers to relatively simple questions have made me look at our current techniques. Also without the answers on the list how can we get the often very stimulating discussion between different contributors. (This also ignores the problem that many questioners no matter how well meaning will fail to post a summary on the list).
As far as time goes I fine that it only takes me three or four minutes to sort out potentially useful items, particularly if good subject headings used. It would take much longer to send individual messages for summaries of questions I am interested in.
My vote for what its worth is to keep the discussions open and active
Ian
} OK, so Gary Login's post was not commercially based. I'm sorry. I was } going to send the question to Nestor Zaluzec only but I got distracted and } sent it to the list by mistake. However, I think it does bear some further } comment and/or discussion. } This list is getting very large and there are now many cases of messages } that should not be sent to all subscribers without there being some } indication that a large number of them want to see the responses. It } really shouldnt have been posted to the list at all, and my reasoning for } that is below. } } Net etiquette (or Netiquette as it is typically known ) says that if there } is a question in a mailing list or newsgroup then the correct response is } to send a message to the original poster. Often a poster will say "private } replies please, if there is enough interest I will summarize to the } net/list". This should be implicit in ALL posts. In this manner if the } poster receives many replies, and there is much duplication in the replies, } then any summary can be edited by the original poster before sending the } information out to everyone on the list/net. People wishing copies of the } replies also send requests to the original poster. } If the original poster geets say five to ten "me too" requests for the } information } You may ask why should the original poster have to do this editing and } summarizing work? Well they are getting the free info courtesy of others } so it isnt exactly a large price to pay. } } Maybe these instructions should be part of the files that gets sent to new } subscribers.
Ian Hallett HortResearch Mt Albert Research Centre Private Bag 92 169 Auckland, New Zealand Fax 64-9-815 4201 Telephone 64-9-849 3660
I think that there is a lot of truth to both sides of this issue. One point that I would like to make is that sometimes responses to (say) a commercial posting are worse than the original! On a few newsgroups (not this one) I have recently seen } 30 responses to a posting with a subject header:
"WOW! This REALLY works! Download it NOW! - money.zip"
The responses to this email (which apparently was } 7000 lines wrong) are now more annoying than the original !
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Why don't we drop the nerdy little cyberjargon? After all we on this listserver are all technogurus of the highest order and can afford to speak English, or other real language, aren't we?
Joiner Cartwright, Jr.
P.S. Of course, you all do realize that I hava a bigger hard disk than rest of you compuwimps, don't you?
Does anyone know of any Macintosh apps. which can be used for plotting orientation matrices/Euler angles (from backscatter Kikuchi diffraction patterns) into inverse pole figures? I can do the regular pole figures already using BKD4.4 by Stuart Wright, but wish to see if any trends become more apparent in the inverse figs. I'm aware of the software available from TexSEM Labs for Windows and SGI, as well as popLA (for IBM compats.) from Los Alamos, but would prefer something for the Mac.
Although most of us would like to see replies to the listserver, I'm sure many will come directly to me. So I will post a summary if there is interest.
Thanks!
Bob Keller NIST Materials Reliability Div. Boulder, CO
Now, if all of you had followed the rules, then I would have been able to post a concise summary saying that I had 12 replies to my message about this and 10 of them were in disagreement and two were in agreement and so it seems that on this list Netiquette is out the window. And we have our usual free-for-all. Sorry, since we should be democratic I guess I'll bow to the majority:-).
BTW. These were not Netiquette a la John Mansfield, rules they were Internet rules formulated in the early days of the net.
John Mansfield North Campus Electron Microbeam Analysis Laboratory 413 SRB, University of Michigan 2455 Hayward, Ann Arbor MI 48109-2143 Phone: (313)936-3352 FAX (313)936-3352 jfmjfm-at-engin.umich.edu, John.F.Mansfield-at-umich.edu or jfmjfm-at-umich.edu they all reach me! URL: http://wwwpersonal.engin.umich.edu/~jfmjfm/jfmjfm.html
I agree with Joiner Cartwright, Jr. that we should avoid jargon whenever possible. It takes a few more key strokes but it will be better communicated to a larger audience. OR, if there are certain well established abbreviations, perhaps they could be posted in list of FAQ's (frequently asked questions). Now, about that hard disk .....
} } } } } } } } } } } } } } } } } }
} Why don't we drop the nerdy little cyberjargon? After all we on this } listserver are all technogurus of the highest order and can afford to speak } English, or other real language, aren't we? } } Joiner Cartwright, Jr. } } } P.S. Of course, you all do realize that I hava a bigger hard disk than rest } of you compuwimps, don't you?
############################################################################# John J. Bozzola, Director Center for Electron Microscopy 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 #############################################################################
Friday -- November 10th, 1995 Joe Hanon's Restaurant 2430 Old Dorsett Drive St. Louis, Mo.
This combined meeting promises to be both informative and practical with discussions of modern, analytical technologies in light microscopy, electron microscopy and molecular biology. Our societies have assembled top-notch researchers and speakers in these areas. The physical setting should be excellent and convenient to reach. Plan to attend this exciting meeting.
9 AM Registration & introductory comments
9:30 Richard L.. Ornberg, Monsanto Company Energy Filtered Imaging with a Light Microscope
10:30 Break -- Refreshments by Electron Microscopy Sciences
11 Jon J. McCarthy, NORAN Instruments New and Emerging Technology for X-ray Spectroscopy in Microanalysis
12 Lunch
1 PM Nestor Zaluzec, Argonne National Laboratory Computers in Mcroscopy, Connectivity and Telepresence Microscopy
3 Susan Wente, Washington University Epitope Labeling
4 Business Meetings
HOTELS, MOTELS:
Drury Westport Inn Single rate $64.00 + tax I-270 & Dorsett Road Includes complimentary breakfast 314-576-9966
Best Western Park Hotel Single rate $69.00 + tax 2434 Old Dorsett 314-291-8700
DIRECTIONS:
From I-70 West and East: Turn onto I-270 south. Exit east at Dorsett Road Make left at first stop light.
From I-40 West and East: Turn onto I-270 north Exit east at Dorsett Rd. Make left at first stop light.
############################################################################# John J. Bozzola, Director Center for Electron Microscopy 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 studying the translocation of the steroid receptor from the cytoplasm to the nucleus after treatment with agonists, using fibroblast L929 cells. The steroid receptor is stained using a specific primary antibody followed by a secondary FITC labelled antibody. To make a long story short, after the treatment with agonists, the fluorescence in the nucleus increases, and wityh my eyes at the microscope this effect is very evident. Now my wish is to use the NIH image programme to quantify the fuorescence in the cytoplasm and in the nucleus, NOT to obtain the absolute quantification of the receptor but to demonstrate and quantify the translocation, i.e., with different agonists. I am using a CCD-72 series camera (DAGE-MTI) to transmit the images from the microscope to the computer.
NOW, I HAVE TWO PROBLEMS:
- FIRST: when I play with the various control functions of the camere, such as "black level" and "gain", both the image I see on the screen and the pixel values change, of course. The problem is that some changes may actually decrase the difference between quantification of fluorescence in the cytoplasm and in the nucleus, while other increses it. For example, the ratio between the average pixel value in the nucleus and the average pixel value in the cytoplasm can range from 2 to 10 IN THE SAME CELLS, just moving the control functions (and it is not related to fading). Can you give me any advice on how to use the control functions?
- I have also a big differences between different experiments or even slides in the same experiment, may be due to the fact that even little differences in the thickness of mounting media can change quantification of fluorescence. In this case, keeping fixed the control functions, I still can get the difference between cytoplasm and nucleus, but the absolute average pixel values in the two compartments may vary from slides to slide, i.e., 20 and 40 in one, 100 and 200 in other. Do you have any advice to solve this problem??
Sorry if the questions required a lot of space
Thanks in advance
Carmine M. Pariante Dep. of Psychiatry Emory University School of Medicine Atlanta, GA, USA phone: (404)-727-8261 fax: (404)-727-3233 cparian-at-emory.edu
Dear colleagues: Can we find another name for this thread, please? There might actually be folks out there with an interest in microwave techniques, and I don't want to wade through all this "Netiquette" stuff to find them. Steven Slap, Energy Beam Sciences really interested in microwaves & EM
I am replying to the list, as I have received several "me, too" requests. There are several options available: a gold on carbon test specimen with particle sizes 5nm to 150nm; an aluminum-tungsten dendrite specimen, designed for use in the 25nm-75nm range; a cross grating replica with 19.7 lines/mm; etc. Please contact me directly for details. Steven E. Slap, Vice President
Dra Patricia Pons was asking for a green chromogen to develop PAP reactions. We found one accidentally which might work. We use the Histomark True Blue substrate and then fix the section in Bouin's fix-the True Blue turns green. You can then was the slide in water or buffer to get rid of the Bouin's. The one thing you have to watch is that TrueBlue in our hands is solvent soluble. We air dry the sections before coverslipping.
Mark Elliott, UBC-Pulmonary Research Lab Vancouver, BC Canada
Message-Id: {199511062228.QAA04132-at-bcm.tmc.edu} X-Sender: joiner-at-bcm.tmc.edu X-Mailer: Windows Eudora Version 2.1.1 Mime-Version: 1.0 Content-Type: text/plain; charset="us-ascii"
At 01:33 PM 11/6/95 EST, "ENERGY BEAM SCIENCES, INC" wrote:
} Dear colleagues: } Can we find another name for this thread, please? There might actually be folks } out there with an interest in microwave techniques, and I don't want to wade } through all this "Netiquette" stuff to find them. } Steven Slap, Energy Beam Sciences } really interested in microwaves & EM } **************** It seems that maybe we have a consensus and can drop it.
We are undertaking to analyze an unusual mineral which contains Mo, As, Ni, Fe, S and apparent C. I am aware of many of the pitfalls of analyzing C (peak shape variability, sample cleanliness, shallow depth of C x-ray production, etc.), but would like to find a better standard than diamond. Can anyone point me towards some heavy element carbides (e.g., Mo-C, Nb-C)?
Please respond via personal email; I will summarize responses to the group if there is sufficient interest. (No, JFM did not tell me to do this ;-) ).
Thanks in advance.
Carl
Carl Henderson Electron Microbeam Analysis Laboratory-Central Campus University of Michigan 2005 C.C. Little Bldg. Ann Arbor, MI 48109-1063 USA -------------------------------- (313) 936-1550 (voice) (313) 763-4690 (FAX) chender-at-umich.edu (e-mail) --------------------------------
Do any of you have a current favorite method for enhancement (silver or whatever) of 5 to 10 nm colloidal gold for light microscopy? The person who is asking will be labelling plant goop on a slide, not sections of embedded material. And is darkfield the best way to view? I haven't had to look for gold with LM before - I rather presume silver enhancement would be the same as for TEM, but more...?
Thanks for any hints!
Tina
***************************************** Tina (Weatherby) Carvalho * Biological Electron Microscope Facility * University of Hawaii * (808) 956-6251 * tina-at-ahi.pbrc.hawaii.edu * http://www.pbrc.hawaii.edu/bemf/ * *****************************************
Some years ago, I used to get a corrosion vascular casting resin for SEM observation called Mercox direct from the manufacturers/ distributors in Tokyo, Japan. My stock has finally run out.
The manufacturers were origionally called Japan Vilene Ink & Chemical Coy and later just Vilene Hospital.
However, I don't have a current address. Does anyone have an address where I can get Mercox from now, either from Japan, or elsewhere? Current price would also be useful if you have it.
Please send replies direct to Bren.Gannon-at-flinders.edu.au
(I will post the addresses to the list, if anyone else indicates interest)
Bren. Bren Gannon Microcirculation & Lymphology Lab Anatomy & Histology Dept., Flinders Univ Medical School GPO Box 2100, Adelaide S.A 5001 Australia Voice (61-8)-204-4183; Fax (61-8)-277-0085 E-Mail Bren.Gannon-at-flinders.edu.au
I think it's time to draw the line. Let's get back to Microscopy & Microanalysis. I for one have had my fill of the last few days of postings about Net etiquette.
You should all remember when I see something that I feel crosses the line, then I send a private note to the author. You are all welcome to do the same, but we don't need copies sent to everyone on the list.
I have NO problems with people responding to questions on the listserver, that is one of our defined operating modes. If the answer involves a product that is for sale, that is also acceptable, so long as the answer is in direct response to a posted question. However, I expect the individual to be brief and to the point. If the posting becomes a commerical then I will politely inform the author. If it goes to far, well......
Remember these postings give everyone the benefit of OUR collective knowledge. This includes both public domain, and commerical material. Let's face it folks there is a lot of information out there that most of us do not know exists, or have forgotten about.
Also , if some kind soul, volunteers to collect a group of related messages and correlate them into one document that is equally acceptable way of getting the information across to this group. There are pluses and minuses to the method. I have no preference.
Please DONOT post any agreements/disagreements we have had far too much bandwidth on the topic. I think all bases have been covered.
Regarding replies to queries posted to the list, the consensus appears to be that all messages should appear on the list. I support this view.
There is however a little technical twist. A number of replies to any posting are "private" by default. Some mailing systems (like the one used by my company) will send the reply directly to the original sender, not to the list. As a result, some queries seem to get no answer or messages appear in reply to a reply that never showed up on the list in the first place.
Such a system is easy to spot: it will usually display the original sender's address in the mailbox directory and in any case put the original sender's address in the "To:" field when a REPLY command is issued. If you are in this case, either replace the address with that of the list in the "To:" field or add a CC: to the list when replying. Also, in some cases a configuration change may do the trick permanently. Check your local SysOp for details.
Regards, MICHEL
Michel Deschuyteneer 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 8113
In light microscopy, Colloidal Gold as well as Silver grain stains are best viewed in Reflected Polarized light. This requires an Epi-Illuminator with a Polarizer and Analyzer crossed to one another. If your microscope is equipped with a Fluorescence Illuminator you just need a set of filters, with a half-mirror (50%) in place of the Dichroic mirror and polarizer and analyzer replacing the exciter and barriers respectively. While Darkfield will identify stained regions, polarized light is most suitable.
Good Luck!
Lawrence Kordon Nikon, Inc. Columbia, Maryland (GO BALTIMORE BROWN!)
In light microscopy, Colloidal Gold as well as Silver grain stains are best viewed in Reflected Polarized light. This requires an Epi-Illuminator with a Polarizer and Analyzer crossed to one another. If your microscope is equipped with a Fluorescence Illuminator you just need a set of filters, with a half-mirror (50%) in place of the Dichroic mirror and polarizer and analyzer replacing the exciter and barriers respectively. While Darkfield will identify stained regions, polarized light is most suitable.
Good Luck!
Lawrence Kordon Nikon, Inc. Columbia, Maryland (GO BALTIMORE BROWN!)
Hi, I know someone in Europe who is urgently in need of an Ion - Mill. No particular preference for model or type. If you know of an available used mill could you please let me know ?
Thanks
lucio *********************************************************************** Lucio Mule'Stagno Physics Dept & Center for molecular electronics Univ. of Missouri- St.Louis tel 314 - 516 5933 e- mail LUCIOM-at-NEWTON.UMSL.EDU
"...if you can judge a wise man by the color of his skin, than mister you're a better man than I .. " Aerosmith *************************************************************************
I disagree with John's view that replies should be routinely posted privately. I learn a lot by scanning the replies, often when I've not even noticed the original query. Provided that a subject field is entered, it's very easy just to delete items which one doesn't want to read.
Ritchie
Ritchie Sims phone: 61 9 3737599 ext 7713 Department of Geology fax: 61 9 3737435 University of Auckland Private Bag 92019 Auckland New Zealand
I fully agree with mr. Sims, especially with what he states about the subject field. Possibly it is even better if every replier uses (exactly) the same subject description (RE: {original subject header} ), so that the mail sorter can sort them together (and they can be deleted together if not of interest).
Adriaan Houtsmuller
* Adriaan Houtsmuller, * Research School for Pathophysiology * of Growth and Differentiation, * Department of Pathology, * Erasmus University Rotterdam (EUR), * P.O. BOX 1738, * 3000 DR Rotterdam, * The Netherlands, * Phone +31 10 408 7499 * Fax +31 10 436 6660
We have been able to locate two standard materials which may work: MO2C, available as a powder (44um particles or smaller) NbC, available as a fine powder (5um particles approx) Please contact me directly for more information. Best regards Steven Slap, Vice-President
This was always available from Ladd...however I'm not sure if they are still in business.
Several years ago, in an attempt to find a lower cost alternative to Mercox, I experimented with some of the acrylic casting resins that are available at hobby shops. These are for making casts of interesting objects, insects, etc. For my purpose (rat carotid), they worked well. Unfortunately, I can't recall the name of the products that I tried. I do remember though that they were but a fraction of the cost of Mercox. Hope this is of some use. ======W. L. Steffens, Ph.D======== ==Dept. of Veterinary Pathology=== ==College of Veterinary Medicine== ======University of Georgia======= =========Athens, GA 30602========= ====STEFFENS.B-at-CALC.VET.UGA.EDU=== =======Voice: (706) 542-5536====== ========FAX: (706) 542=5828======
Although eip-polarized light is the fastest way to look directly at gold labeled sections it is certainly not the most economical if you are not fully equipped to begin with.
SIlver enhancement is a simple alternative and works the same as when used for EM. The silver solution is reacted with the gold and then fixed using a solution similar to photographic fixers. As there is no worry about grain size any mordanting steps can be omitted. A simple kit which has worked well for LM in my hands (J. Cell Biol. 1988 106:279-288) is a kit from Amersham called "IntenSe". The silver solution is not light sensitive so it is possible to check the cells or sections with the light microscope before fixing, repeating the reaction until a signal is detected. Detection is easy using transmitted light or phase contrast, the signal appears as black precipitates.
Another alternative is to use enzyme histochemistry, where the signal is detected as a colored reaction product (HRP, Alk.Phos. etc).
I have placed a small discussion on this subject on the WWW - URL http://info.med.yale.edu/cellimg/Cryo-LM.html if you are interested.
The standard procedure for enhancing colloidal gold particles with a silver enhancement kit is: 1) Wash section very thoroughly with distilled or deionized water 2) Mix 3 drops each of initiating solution and enhancing solution in a clean test tube 3) Add several drops of the mixture to the slide and monitor development of silver stain periodically under the microscope at room temperature. Do not expose slide to high intensity light for prolonged periods. Staining is typically complete when stained areas have a brown-black stain. Stop the reaction before non-specific background appears by thorough washings in distilled water. 4) To stop the reaction, wash thoroughly with water. If development is not complete, use fresh solutions and continue silver enhancement. Counterstain as desired.
Note that this procedure was written for our own BioSite silver enhancement kit, but should be applicable to most high quality commercially available kits.
These instructions can be found at our WWW site as part of a complete instruction brochure on use of BioSite colloidal gold reagents. (http://www.mwrn.com/ebs/ebs.htm) Steven E. Slap, Vice-President
Tardis, Los Alamos, New Mexico was asked by a client to combine a stack of images each with a limited depth of focus into one image that shows all areas in focus. They did a very nice job and I am encouraging them to add the interface and software that would make this a universal tool.
This is where you come in, is there appreciable interest in something like this?
Another thing of course is can it be made universal and easy to use? Are there applications where it doesn't work? We need feedback and image sets to drive this project forward. The Alpha image set and result can be seen at:
http://www.rust.net/~jlafeber
From what I have seen of image sets (six) at this time the Tardis approach is robust and very, very fast (under 1 minute with a standard Pentium PC).
I am seeking a fixation protocol and staining procedure for cadmium in embryos for the TEM. I am very interested in the histochemical staining procedure and protocol using Benzothiazolylazonapthol derivatives (BTAP's) synthesized by Sumi Y, Muraki T, and Suzuki T (1979, 1980, 1982). BTAP's stain differentially stain cadmium in tissues embedded in paraffin. Any help would be appreciated. Thank-you. MScroggie , university student, Trent University. MSCROGGIE-at-TRENTU.CA
} Aloha, microscopists, } } Do any of you have a current favorite method for enhancement (silver or } whatever) of 5 to 10 nm colloidal gold for light microscopy?
I routinely enhance labelled wholemounts of retina (300 mic thick) with BioCell (UK) silver enhancing kit. Wash labelled, fixed tissue with water before and after enhancing; enhance till visible under LM. View normally, with tissue mounted under glycerol, as dehydrating seems to turn everything black. It is also more sensitive if you use 1nm gold conjugate as the initial label (also available from BioCell). Darkfield is more sensitive, but normal LM is fine and you can see the rest of the tissue.
Diana van Driel Dept Ophthalmology Sydney University Sydney 2006 Australia
I'm working with a group that wishes to make morphological measurements on polymer nanoparticles using electron microscopy (SEM or TEM). The nanoparticles will likely be polyvinylpyridine, polycyanoacrylate, possibly polymethylmethacrylate. The nanoparticles should be in the range of 100-200nm, but they may well be bigger. They would like to see the whole range of particles and be able to report average particle diameter and the whole range of particle sizes (size polydispersity).
Previous attempts, by placing drops of particles suspended a solvent on a SEM mount or TEM grid, have resulted in "clumps" of particles that are difficult to measure.
Can anyone recommend a prodecure that might work for this situation. I have heard of (but am unfamiliar with) aerosol procedures, will that work?
Also, can anyone recommend a few useful general references on electron microscopy of polymers. I have "Polymer Microscopy" by L. Sawyer & D. Grubb on order now.
Thanks. Owen P. Mills Michigan Technological University Metallurgical & Materials Engineering Rm 512 MME Building Houghton, MI 49931 906-487-2002 906-487-2934 FAX opmills-at-mtu.edu
I am working on an undergraduate thesis to improve the resolution of xray detectors for EDS systems, likely using a cryogenic detector. Our goal is to achieve a resolution of about 10eV (for Mn K alpha), while retaining reasonable count rates (allowing faster processing then wavelength dispersive spectrometry). I would be interested in hearing about specific applications for such a system (such as trace element analysis or light element detection) and about the specific shortfalls of current EDS systems with less resolution (such as two peaks that you would really like to resolve but can't). If convenient, any references to literature would be appreciated.
Thank you for your time.
Don Morgan Queen's University Kingston, ON (613) 530-3506
} I am working on an undergraduate thesis to improve the resolution of xray } detectors for EDS systems,.................. references appreciated..
Don,
You should get a copy of the Proceedings of the 29th Annual Conference of the MicroBeam Analysis Society, held in BreckenRidge Co, Aug. 6-11, 1995. Editor: E. S. Etz by VCH Publishers.
There were several papers on the topic of ultra high energy resolution EDS detectors there, and plenty of references in the papers therein.
You might also consider alternate means of parallel detection of x-rays. such as combining multilayer x-ray optics and multichannel position sensitive detectors and/or CCD arrays. It brings in a totally different bag of worms, but nevertheless is an alternate approach.
You should for example look up the work of Dave Wittry and colleagues. Try in the J. of Applied Phys. ~ 1990-1994 there are several papers by him on the topic of PolyChromatic Diffraction Spectrometers (i.e. Parallel detectors for X-ray Microanalysis). as well as one in Ultramicroscopy circa 1988 or 89 entitled "Two Dimensional CCD Arrays as Parallel Detectors in Electron Energy Loss and X-Ray Wavelength Dispersive Spectroscopy" by Strauss & Zaluzec (sorry but I don't have the exact reference on that one handy).
Remember it is not just energy resolution that is the important factor. You also need good geometrical collection efficiency, small size (so that your detector will fit in your microanalysis instrument) and the ultimate controlling factor a reasonable price tag.
While in Germany a few years ago, a colleague bought a xylene-based medium called Melanol for making permanent (or at least semi-permanent) mounts. She has just about run out but can't find a supplier to buy more. Is there a new supplier of this mountant? If not, can anyone advise re. a suitable, less toxic substitute?
TIA, Rosemary White
____________________________________________________________ Rosemary White __ / Department of Ecology _/ \__/ \ and Evolutionary Biology / \ Monash University / Australia \ Clayton, Victoria 3168 \ ____ / phone 61-3-9905 5670 \_/ \_*_/ fax 61-3-9905 5613 __ email r.g.white-at-sci.monash.edu.au \/
Dear Fellow Microscopists, I=B4m looking for EM micrographs of periodical (biological) objects (particles, macro molecules), suitable for course introduction to FFT and/or electron diffraction). Is there any internet (www, ftp) source for such demo-images?
Thanks for any response, -Dietmar-
+++ Dietmar Reiter {dietmar.reiter-at-uibk.ac.at} ++++++++++++++++ +++ Dept. of Zoology and Limnology, University of Innsbruck ++++ +++ Technikerstrasse 25, A - 6020 Innsbruck, Austria +++++++++ +++ ph: (+43)-512-507-6170 (-6161), fax: ..-507-2930 (-2957) +++
... If I would have had more time, I would have written a shorter e-mail ...
A paper ["Quantification of particle sizes with metal replication under standard freeze-etching conditions: a gold ball standard for calibrating shadow widths was used to measure freeze-etched globular proteins" Microscopy research and Technique 32 (1995) 312-329.] was just published that employs 45deg. unidirectional Pt-C replication to measure particles. This paper will give you insight into the problems of replication and how to avoid the pitfalls---- at the end of the paper is a vertical replication method which (Fig. 17b) is the most powerful and reliable of the replication methods. This method employs a small metal coating correction to achieve the actual particle size (see ref in paper). This method would allow you to measure the asymmetry of the particles as well. Another method which is more readily available to you but can not help you in the 1-2 nm size range is a negative staining technique. For this method you would need to make up a 2% solution of Uranyl acetate in a tin foil covered bottle (light sensitive) and filter thru a 0.22 micron filter before use! You would need 10-12 nm thick carbon films covered 300 mesh grids. Use a 10 microliter drop of your particles in ethanol, propyl or isoppropyl alcohol or some solvent compatible with water ---- and put it on the carbon film for a minute. Remove the excess alcohol from the edge of the grid with torn Whatman #1 filter paper. Apply 5-10 microliters of stain for a minute and remove with the filter paper method and repeat the procedure! Dry the grids from the edge with torn Whatman #1 filter paper and then look at your grids in the TEM. You will want to measure the white particles you see surrounded by dark stain.
George C. Ruben Dept. Biological Sciences Dartmouth College Hanover, NH 03755 603-646-2144 603-646-1347 fax George.C.Ruben-at-Dartmouth.EDU
} From: Bernard A Klazema } Another problem we have is : we bought a new SEM (Philips XL30) and the image w } e make must be printed on a Kodak 8600 PS dye sublimation printer. TIFF format. } We like to have an overlay on the images with all the data like U marker and } Mag on it, As far as I know this is only possible by a videoprinter, but is the } re someone who can help me out with some software (Macro in windows?) } We have in house Coral Paint and all the normal software packages from Micro- } soft.
I have done well with the Recorder program that comes with Windows in the Accessory group. It provides macro capability for all Windows applications, not just those with built in macros. It can record mouse actions as well as keystrokes. I have strung several macros together with their own hot keys. Say the macro first selects the text tool, then a location, and perhaps even starts filling in the text. I end that macro there and define another one to finish the task. One might argue that it is no substitute for a good macro tool in the application itself, nor does it allow editing of the recorded macros, but it still can do a lot if one is careful and creative.
I have also made use of the cut and past features of Windows apps when labeling images in PhotoStyler. Since most all text boxes will accept pasted text, I have opened Notepad or Write to compose a set of standard labels and then copied the appropriate starter text (to the Clipboard) for pasting into the image. I then customize it for the particular image or mag or exposure, etc. It has saved me a lot of time. ---------------------------------------------------- Warren E. Straszheim 270 MD Ames Lab/ISU, Ames IA, 50011 Phone: 515-294-8187 FAX: 515-294-3091 E-Mail: wes-at-ameslab.gov (or: wesaia-at-iastate.edu)
coal characterization and processing electron microscopy, x-ray analysis, image analysis computer applications
} Anyone got a protocol for the preparation of the compound eyes of insects } (Drosophila) for scanning electron microscopy? } } If so, would you be willing to share it? } Sure.
We collect insects from the fluorescence light in our kitchen for scanning electron microscopy examination. These are nicely dried without collasping. The bugs (eyes and all) look great and several have of the images obtained from these specimens have been used in advertisement.
A list of images on the WWW is in the "Guide to Microscopy and Microanalysis on the Internet" at the MicroWorld Resources and News WWW site. The images are listed by subject. Each techniques category also has sites with images marked.
MicroWorld Resources and News is at: http://www.mwrn.com/
Please check the information at each site to determine if there are any restritions on the use of the images. If you have any problems, please send me a message.
In my prior life as a chemical engineer, I was responsible for a colloidal silica unit. Whenever questions were raised about particle size distribution, we would walk a sample over to our friendly electron microscopist for analysis.
His secret was to perform a series of dilutions to very dilute levels of Si. My recollection is that he did a series of three or four 1:100 dilutions before drying a drop on a coated grid. This created a single particle thick film of these normally sticky SiOx particles.
We were routinely dealing with discrete particle sizes from 6 to 150 nm.
Although it was 15+ years ago, I could dig up his name and number if you would like to give him a call.
Good luck, Joe Tabeling Delaware Diamond Knives 800-222-5143
Fellow microscopists We have a couple of EMs with digital imaging. We also have a lexmark 1200 dpi laser printer. I need a software package which will allow me to print batches of tiff files without loading the individual files first. It would also be nice to be able to decide how many images to have per page. I use ibmpc and mac on a novell 3.12 network. Any help appreciated.
Chris
Chris Gilpin Biological Sciences E.M. Unit g452 Stopford Building Manchester University Oxford Road Manchester M13 9PT U.K. 0161 2755170 phone 0161 2755171 fax
We are finishing are shopping for a new SEM. Since I have not used a new digital SEM before, I have some basic questions.
(1) Do we still need to purchase a Polaroid camera and high resolution CRT system, or are people comfortable going completely digital? It looks as if we omit the camera, we would have enough left over for a good dye-sub printer and laser printer.
(2) Frame capture boards. Is a resolution of approx. 1K x 1K enough or is the extra $$ for a 2K x 2K necessary?
We currently have a Zeiss TEM and are considering the new LEO SEMs. Any comments on this new merger of Zeiss and Leica electron optics?
Thank you for your help. As we are a small college, our decision will probably be around for a long time...
Page Owen Connecticut College Department of Botany New London, CT 06320 (203) 439-2147
Has anyone heard of an increased incidence of cataracts in electron microscopists ??? I have just been told that I have cataracts developing and that it is unusual to see them in someone my age. I can't help but wonder if my choice of profession had anything to do with it.
Thanks for any info.
Barbara Hartman Schering-Plough Research Institute Barbara.Hartman-at-Schering-Plough.sprint.com
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Dear Microscopists -
Does anyone use Forier Transform Infrared Microspectroscopy ("FTIR") on biological materials? I would be interested in learning about this technique and hearing of your experiences with it. This is a new method of identifying compounds with microscopic resolution.
Joiner Cartwright, Jr., Ph.D. Director, Electron Microscopy tel.: (713)798-4658 Department of Pathology, Rm.286-A FAX: (713)798-3945 Baylor College of Medicine Internet: joiner-at-bcm.tmc.edu One Baylor Plaza Compuserve: 71555,1206 Houston, Texas 77030 U.S.A.
The latest volume edited by M. A. Hayat, "Immunogold-Silver Staining: Principles, Methods and Applications" (CRC Press, Boca Raton, FL, 1995) has many articles on methods of silver enhancement. The earlier series, "Colloidal Gold: Principles, Methods and Applications" (M. A. Hayat, Ed.; Academic Press, San Diego, CA: Vols. 1, 2 (1989) and Vol. 3 (1991) have some useful info as well.
Richard D. Powell Nanoprobes, Inc (We sell silver enhancers too). URL: http://www.lihti.org/research/ecodev/incubten/nano/home.html
} Do any of you have a current favorite method for enhancement (silver or } whatever) of 5 to 10 nm colloidal gold for light microscopy? The person } who is asking will be labelling plant goop on a slide, not sections of } embedded material. And is darkfield the best way to view? I haven't } had to look for gold with LM before - I rather presume silver } enhancement would be the same as for TEM, but more...?
We do this type of analysis by cryoTEM of vitrified thin liquid films of dispersed particles. We collect the images on a slow-scan CCD camera using Gatan's DigitalMicrograph and do image analysis using NIH Image. We compute the size distributions using a statistical package called RS/1 (on the VAX or Sun) and fit the data to a lognormal distribution with up to three modes.
We have tried aerosol methods and have not been satisfied with the fraction of isolated particles. We do MUCH better with cryoTEM. The good news is that one can automatically select only isolated spheroidal particles from the images containing some overlapping particles by calculating the circularity of each feature (C=4*pi*area/perimeter**2). For such particles we only keep features with 0.90 } C } 1.05. Verify this yourself by measuring model images of agglomerated circles. By the way, it took me almost a year to develop this capability to the point that I convinced myself that I had done everything correctly. It was not a trivial project.
} Hello; } } I'm working with a group that wishes to make morphological measurements on } polymer nanoparticles using electron microscopy (SEM or TEM). The } nanoparticles will likely be polyvinylpyridine, polycyanoacrylate, possibly } polymethylmethacrylate. The nanoparticles should be in the range of } 100-200nm, but they may well be bigger. They would like to see the whole } range of particles and be able to report average particle diameter and the } whole range of particle sizes (size polydispersity). } } Previous attempts, by placing drops of particles suspended a solvent on a } SEM mount or TEM grid, have resulted in "clumps" of particles that are } difficult to measure. } } Can anyone recommend a prodecure that might work for this situation. I have } heard of (but am unfamiliar with) aerosol procedures, will that work? } } Also, can anyone recommend a few useful general references on electron } microscopy of polymers. I have "Polymer Microscopy" by L. Sawyer & D. Grubb } on order now. } } Thanks. } Owen P. Mills } Michigan Technological University } Metallurgical & Materials Engineering } Rm 512 MME Building } Houghton, MI 49931 } 906-487-2002 } 906-487-2934 FAX } opmills-at-mtu.edu
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Does anyone out there in microscopy land use a software package called "DragonDictate", and are willing to share their thoughts about it? We have some people in our department who are considering such an automated dictation setup. They are impressed with IBM's program, but are curious about DragonDictate. Does anyone here in the Houston, Texas area have it and want to show it off?
Joiner Cartwright, Jr., Ph.D. Director, Electron Microscopy tel.: (713)798-4658 Department of Pathology, Rm.286-A FAX: (713)798-3945 Baylor College of Medicine Internet: joiner-at-bcm.tmc.edu One Baylor Plaza Compuserve: 71555,1206 Houston, Texas 77030 U.S.A.
} To MSA subscribers I am an undergraduate student seeking higher } education in the fields of SEM and TEM. I will be graduating soon } and I am searching for a college that will offer certification in EM. } Can anyone help me in this area? Come to the University of Arizona! We have an outstanding Materials Science and Engineering microscopy facility, and full semester courses on both TEM and SEM.
Of course, there are many other fine public and private universities offering EM classes. Gain access to a World Wide Web browser, and look around. Try doing a web search for "microscopy and education" or some such thing.
Best regards,
Mark T. Biedrzycki Computer Networking Laboratory Materials Science and for Microscopy Education (CNLME) Engineering Department at the University of Arizona, Tucson
We are having a slight problem with our JEOL2000FXII. We have persistent arcing illustrated by a flickering, unstable beam. We have cleaned the gun area several times with little effect. The dark current is stable. We have also tried several filaments. We also have adequate freon. So, we are at the end of our rope.
I would be extremely grateful if anyone would have a suggestion on what to look for to fix this thing. Cleaning procedures that you use would also be helpful, since we hypothesize that is still the problem. Thanks for your assistance. It could save us $$ in a service call.
Jeff --------------------------------------------------------- U U | Jeff Shield | U U | Department of Materials Science and Engineering | U U U U | University of Utah | U U U U | Salt Lake City, UT 84112 | U U U U | 801/581-3179 Fax: 801/581-4816 | UUUUU U | | U U --------------------------------------------------------- UUUUU Of making many books there is no end, and much study wearies the body. -Eccl 12:12
Digital Micrograph-Demo Site is beeing organized by the MSA Education Committee.
The need for such a site is evident. Within the Education Committee of MSA I try to organize such a "digital image data resource" site for all microscopies. At this time, many laboratories offer plenty of images at their web sites but a central directory and specific copyrights are lacking. Everbody, who wants to contribute to a general "MSA Digital Image Resource", please contact me. We will make the images available through the MSA web server. Images may remain in local web resource directories and may only be linked to the general register.
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Microscopists -
Several months ago there was a posting on the Microscopy Listserver concerning a super bad computer virus called the "Good Times" virus that would end civilization as we know it. Almost immediately a number of replies came back debunking it as rumor. Apparently the real virus was the panic and paranoia that it spread. Well, the scare seems to have cropped up again here in the Houston area.
My question is: Is it still considered bogus? Is there any further development on this?
Joiner Cartwright, Jr., Ph.D. Director, Electron Microscopy tel.: (713)798-4658 Department of Pathology, Rm.286-A FAX: (713)798-3945 Baylor College of Medicine Internet: joiner-at-bcm.tmc.edu One Baylor Plaza Compuserve: 71555,1206 Houston, Texas 77030 U.S.A.
In message {Pine.A32.3.91.951107204433.43418E-100000-at-san_marcos.csusm.edu} Kevin Schram writes: } } Heh all! } } Anyone got a protocol for the preparation of the compound eyes of insects } (Drosophila) for scanning electron microscopy? } } If so, would you be willing to share it? } } Contact me at: } } schra001-at-coyote.csusm.edu } } } Thanks in advance!
Kevin,
I've done SEM of compound eyes of fruit flies (sorry, my Latin ain't good) using my cold stage on my Philips 500SEM. The fly bodies were squished onto carbon double stick tape on aluminum pin stubs, dabbed with a bit of carbon paint, oriented so that the eyes were looking upward....(goodbye cruel world). Then sample was placed onto liquid nitrogen pre-cooled cold stage for freezing, quickly inserted into SEM and imaged at 1.5 kV. No metal coating was applied, thus the reason for low kV and no charging is observed. If any frost was initially observed, I'd shut off the beam, bleed a bit of dry nitrogen gas through the chamber, pump vacuum down again and proceed.
Using this method, images were recorded up to about 2,000X of patterns of wild and mutant fly eyes. I proudly call the technique LVLTLRLMLOSEM (low voltage low temperture low resolution low magnification low overhead SEM). It also works well for some delicate plant tissues.
If you do not have access to an SEM with cold stage you could try the ol' fixation, dehydration & critical point drying method, but we tended to get collapse of eye facets using that technique, but you might do well enough to get the information you need.
Hope this helps. If you want to discuss further, contact me off line.
--
Gib Ahlstrand, MMS Newsletter Editor Electron Optical Facility, University of Minnesota, Dept. Plant Pathology 495 Borlaug Hall, St. Paul, MN 55108 (612)625-8249 612-625-9728 FAX, giba-at-puccini.crl.umn.edu
"MICROSCOPY & MICROANALYSIS 96" in Minneapolis, Minnesota, Aug.11-15, 1996
Page- I would advise to spend the money on the 2k x 2k image capture (if you can afford it), you'll be glad you did 2 years from now when 1k x 1k is considered obsolete. And save your money on the Polaroid, it is obsolete. Invest in two printers, a dye-sub for publication, and a 600-1200 dpi laser printer for working copies. Just my opinion. -Mike
On Wed, 8 Nov 1995, T. Page Owen Jr wrote:
} We are finishing are shopping for a new SEM. Since I have not used a new } digital SEM before, I have some basic questions. } } (1) Do we still need to purchase a Polaroid camera and high resolution } CRT system, or are people comfortable going completely digital? It looks } as if we omit the camera, we would have enough left over for a good } dye-sub printer and laser printer. } } (2) Frame capture boards. Is a resolution of approx. 1K x 1K enough or } is the extra $$ for a 2K x 2K necessary? } } We currently have a Zeiss TEM and are considering the new LEO SEMs. Any } comments on this new merger of Zeiss and Leica electron optics? } } Thank you for your help. As we are a small college, our decision will } probably be around for a long time... } } Page Owen } Connecticut College } Department of Botany } New London, CT 06320 } (203) 439-2147 } } tpowe-at-conncoll.edu } }
There is some rule that when something is going well don't change or stop anything.. for several months I had good success immuno- gold labeling using a polyclonal antibody for an integrin antigen. I could get specific labeling in .3 micron sections of bone embedded in LR White (silver intensified gold) for LM or in thin LR White sections for EM. The same specific labeling results occurred in a dozen experiments; antigen absorption of the antibody and deletion of antibody were negative controls. The antibody was specific and dependable. I had to interrupt the project and did not return for a year. Murphy's law takes over. With the same antibody, using aliquots stored at -20 C as per manufacturers instructions as well as aliquots stored at -70 C, I cannot get labeling on sections cut from the original blocks. The antibody still works on pre- embedded tissue culture. On top of this, the original manufacturer went out of business. The antibody produced by a new company with the original protocol does not work. I thought that maybe the LR White blocks after a year further polymerized to reduce antigenicity. I worked up newly embedded tissue, freshly cut sections, and still got no label. I've tried different secondary antibodies and different blocking and different buffers -- no label. A different antibody for a different antigen does specifically label in sections from old blocks or new. So... can an antibody 'die' even if stored properly? Sorry for this long winded tale but with no substitute available commercially for this particular antibody I'd love to know what happened. Thanks for any comments.
Pat Masarachia Bone Biology Merck Research Laboratories West Point, PA 19486
} I am working on an undergraduate thesis to improve the resolution of xray } detectors for EDS systems, likely using a cryogenic detector. Our } goal is to achieve a resolution of about 10eV (for Mn K alpha), while } retaining reasonable count rates (allowing faster processing then } wavelength dispersive spectrometry).
Dear Don, The theoretical limitation to energy resolution from a solid-state detector, like those used in EDS, comes from counting statistics. For an x-ray of ~6 keV to have a resolution of 10 eV, you need to produce 60 elec- tron-hole pairs per eV of energy loss. This gives 360,000 e-h pairs per 6-keV-x-ray, or +-600 counts per measurement. N.b., this assumes each x-ray is fully absorbed, no small background counts occur simultaneously, and none of the other problems with getting 6 keV deposited in the detector for each x-ray occurs. Unless you are going to operate on a different principle from that used in solid-state detectors, your first problem is to find a material with a small enough band gap. Maybe an appropriate metal will serve as a semiconductor at a low enough temperature (} 1 K). Good luck; this seems a very complicated and difficult problem for a thesis--especially an undergrad- uate one. Yours, Bill Tivol
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Greetings,
I wanted to make folks aware of a program for getting kids into science. I have participated in this progam for several years and find it very rewarding. The kids send letters and questions that are fun to read, and most seem very interested in science. If you have just a few hours to spare, please consider becoming a voluteer scientist. (details below)
Thanks,
James
(Forwarded from sci.chem) SCIENCE-BY-MAIL PROGRAM SEEKING VOLUNTEER SCIENTISTS
Science-By-Mail is a pen-pal program from the Museum of Science in Boston, Massachusetts that teams scientists with children in grades 4 through 9. Due to an increase in membership this year we need additional scientists for the 1995-96 program year. All of the scientists in our program act as pen-pal mentors and correspond with up to five groups of 1-4 children or with one classroom of seven groups of 1-4 children. We mail out two activity packets per year (once in December and once in March) for you to review, and then correspond with your pen-pals about the activities in the packets.
The commitment requires approximately 20 hours per program year (November through June). Our topics this year are the Science of Sports and Planetary Science. Volunteers do not need to be experts in the fields of the topics, we provide information on each topic to the scientists, for each activity.
In order to be a Volunteer Scientist you need to have a bachelors degree in a science or technology related field. You also need to have the desire to inspire scientific curiosity in children.
If you are interested in volunteering for Science-By-Mail please call 1-800-729 3300 or 617-589-0437 and ask for Melissa Cotter or Monica Parker. If you get our voice mail just leave your name, phone number and fax number and we will send you information right away. Thank you in advance for your help! ============================================================================== James C. Long Manager/Materials Analysis Lab Electrosource, Inc. 512-445-6606 jlong-at-bga.com
Dear Page: Despite what you have heard, all the SEMs have been digital for at least 10 years, now, at least where it is practical to be digital. They all still go back to analog for the Polaroid, since that is the most practical way to get photographic resolution. Remember that you can only record the resolution that the instrument puts out and recording a 2K by 2K image out of a 800 by 600 digital output will only get you the lower number resolution. By my experience with SEMs and digital imaging systems, you still need the photographic output for the best quality images and the convenience of film and negative. Most SEMs can output at least 2500 lines on the photo CRT. I have yet to see a laser printer or dye-sub printer which truly gives photo quality images when examined closely. The printers are great for "quick and dirty" and save the cost of the Polaroid film when it is not needed, so it is nice to have both, but if the SEM is just a big camera, then you should judge it by the quality of its best output. I would be very reluctant to lose the photographs. Best of luck. Mary
Mary Mager Electron Microscopist Metals and Materials Eng, UBC 309 - 6350 Stores Rd. Vancouver, B.C. V6T 1Z4 CANADA tel: 604-822-5648, fax: 604-822-3619 email: mager-at-unixg.ubc.ca
I'm looking at serial sections of nerves on formvar (1% in chloroform) coated slot copper grids (pretty standard stuff). The grids are acetone and distilled water cleaned several times and coated as per standard (which has worked consistently well for 4 years). For the past few months I've been having considerable trouble with what appeared to be focus problems. It now appears that the film is "shifting". The film is carbon stabilised and clean; we've tried modifying formvar coating preparations, carbon coating and specimen storage, but the film movement is still present. It appears "tight" when on the grid, but appears to shift under the beam (as indicated by what appeared to be focussing problems). Anyone with a suggestion????????????????????????/
} Several months ago there was a posting on the Microscopy Listserver } concerning a super bad computer virus called the "Good Times" virus that } would end civilization as we know it. Almost immediately a number of replies } came back debunking it as rumor. Apparently the real virus was the panic and } paranoia that it spread. Well, the scare seems to have cropped up again here } in the Houston area. } } My question is: Is it still considered bogus? Is there any further } development on this?
Info from our virus experts is that it is not possible to spread by email unless there is an attached file which is actually run. Anyone doing this would be asking for trouble. The conclusion is that Good Times is not for real, but since we have had recent severe problems, I am interested in information. The problems have been with MOnkey and MonkeyB viruses which are very tricky. cheers jjm Professor John J. Millar Department of Applied Physics and Electron Microscope Unit RMIT, GPO Box 2476V, MELBOURNE AUSTRALIA 3001 +613 9660 2602 fax 613 9660 5290 email jjmill-at-rmit.edu.au
} Greetings, } } We are having a slight problem with our JEOL2000FXII. We have persistent } arcing illustrated by a flickering, unstable beam. We have cleaned the gun } area several times with little effect. The dark current is stable. We have } also tried several filaments. We also have adequate freon. So, we are at } the end of our rope. } } I would be extremely grateful if anyone would have a suggestion on what to } look for to fix this thing. Cleaning procedures that you use would also be } helpful, since we hypothesize that is still the problem. Thanks for your } assistance. It could save us $$ in a service call. } } Jeff } --------------------------------------------------------- U U } | Jeff Shield | U U } | Department of Materials Science and Engineering | U U U U } | University of Utah | U U U U } | Salt Lake City, UT 84112 | U U U U } | 801/581-3179 Fax: 801/581-4816 | UUUUU U } | | U U } --------------------------------------------------------- UUUUU } Of making many books there is no end, and much study wearies the body. } -Eccl 12:12
Jeff,
your microscope is not the only 2000FX or FXII to suffer this problem. We have one of each in our labs, both suffered this flicker in illumination and I know of at least 3 others in Australia. In our case the cermic insulators in the gun chamber had suffered numerous dicharges which left tracks. These tracks facilitated further discharges and the whole problem snowballed. We tried cleaning the insulator from the 2000FX but with no success. In July 1993 we installed a new insulator (at great expense). When the 2000FXII developed the same problem in October 1994 we opted to convert to SF6 (from freon) as the HT tank and gun insulation gas. This involved replacing the entire gun assembly and overhauling the HT tank to make it compatible with SF6. This was VERY expensive.
We have had no repeat of the flicker problem so far. We intend performing an overhaul of each gun assembly bi-annually in the hope that the discharges can be avoided in future.
Mark Blackford TEM Group Materials Division, Ansto PMB 1, Menai, N.S.W. Australia 2234
-- [ From: Charles A. Garber, Ph. D. * EMC.Ver #2.10P ] --
On November 7, Bernard A. Klazema asked about the thin sectioning of "glass filled materials". He suggested this would result in the "ruining" of a diamond knife. His samples are impact modified glass fiber reinforced plastics.
Well, he is half right and half wrong. With practice one can very definitely do ultramicrotomy on such samples without "ruining" the diamond knife. However, one has to appreciate the following:
a) The "wear and tear" put on the diamond knife is considerable, with more "wear and tear" put on by the inexperienced ultramicrotomist, and less, by the experienced ultramicrotomist. But no matter what is the experience, the point is that this kind of microtomy is going to put "wear" on the knife at a rate far greater than that what would be encountered for straight soft tissue samples.
The depreciation of the diamond knife is a significant cost factor in the conduct of this type of work. In our own laboratory, we can usually get 15-50 samples of this type out of each knife, depending on the percentage loading of the glass fibers, and yes, the experience of the person doing the work. But it can be done, provided one is willing to pay the cost associated with the knife's depreciation.
b) We recommend a "materials science" diamond knife (of the type offered by our firm because we do not know of anyone else offering such a product), one that is basically the same "angle" as a life science diamond knife. We do not like the idea of using a "blunter" angle, to get longer knife life because the possibilities of compression effects increase greatly and also therefore various artifacts from the cutting. We have found that the distortion that results, even when done cryo, of these kinds of samples, with a blunter angle knife, caused round rubber modifier particles to come out looking like elipses, oriented in the direction of the knife.
c) The materials science knives supplied by our firm are not "striation free", they do contain, at the edge a small number (e.g. not more than a few) of fine striations. Such knives would not be acceptable for typical life science work. But this small population of fine striations that is there to begin with is smaller than the much larger number of striations put into the knife edge after taking the first slice on a glass fiber filled plastic.
So my point is why pay more for the "perfect" knife when one with a few striations to begin with will do just as well? And at the same time, a substantial amount of money can be saved relative to the price paid for the typical "life science" knife (sans striations).
d) For visualizing the impact modifiers, osmium tetroxide or in the case of acrylic modified sytems, ruthenium tetroxide, can be used to demonstrate the structure and morphology of such systems, including the spatial arrangement of the glass fibers.
Hope these comments will be useful.
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: GVKM07A-at-prodigy.com West Chester, PA 19381-0656 USA Customer Service: SpiSupp-at-aol.com
Jeff, You are most probably correct, that is, the gun chamber is contaminated. Find out at what accelerating voltage the beam becomes stable, if it is a little less than 200kV the gun chamber is probably the problem. Also if you monitor the gun chamber vacuum any HT instabilities will coincide with variations in the gun vacuum. This will give you an idea whether the HT problem is in the gun or the HT tank. Also you can isolate the HT tank from the gun and bring up the HT to 200kV. By monitoring the CH output from the HT tank with a CRO you should be able to monitor the high voltage ripple waveform. This waveform is the ripple of the HT at whatever kV the HT is applied at and if any instabilities are present they will be seen as sharp rapid spikes within this waveform. The typical waveform pattern is {5mVp-p AC -at- 20ms/cm. Be carefull not to blow up your CRO. If any large discharges occur in the HT it will kill your input pre-amps to the CRO. You say the beam current is stable but the flickering is evident. The beam current is definitely unstable it is the meter that cannot detect any small rapid variations. Have you tried conditioning the HT above 200kV for a short period of time and see whether the beam is stable after this. I still rekon the problem is with contamination. Especially since you say you have cleaned the gun chamber several times. Every time you have cleaned this region no matter how careful you are you will have left behind some sort of contamination. I never use any polish except for the wenelt assembly and anode only when definite contamination is present. All other parts are wiped clean with a lint free cloth soaked in liquid freon. All polished parts must be ultrasonically cleaned in freon and then baked before re-assembly. Even with this procedure the beam instabilities you are experiencing I to have seen at 400kV though. The routine maintenance procedure to eliminate this slight gun chamber beam instability is to create a HT glow discharge in the gun chamber. This is done by raising the gun chamber vacuum to 210uA on the pirani gauge and applying the HT at (I do not know for the 200kV machines) 150kV for the 4000EX series machines. You have to overide a lot of protection circuits to be able to achieve this and experience and care must be taken into account. If you need help I have a complete discription explaining this procedure but first check with Jeol to see if it can be done for your machine and at what kV is best. The procedure is done with an inert gas, usually Nitrogen, and what happens is a glow discharge which moves all the contamination around. About 10% of the contamination is removed by the rotary pump and the rest is moved out the way of the general beam line. Within time the procedure is needed again. For the 400kV machines this is routine about every 6 months. In summary I can guess your machine was O.K. untill you opened the gun for some other reason. And once closed and maybe some time down the track this instability happened. I have several questions that will help to pin point the problem and maybe it is best you e-mail me direct. What is the gun vacuum? What kV does the beam become stable? Have you tried HT conditioning? Have you cleaned the upper and lower anodes? Have you isolated the probelm to the Gun region by monitoring the Gun chamber vacuum?
Regards, David Dryden School of Physics University of Melbourne Parkville, VIC. Australia, 3052. djd-at-electron.ph.unimelb.edu.au http://www.ph.unimelb.edu.au/~djd/diff-home.html
This is a summary of the responses to my question of two weeks ago, which was: "How do I keep commercial absolute ethanol dry once the container has been opened?"
Firstly, to those who replied my thanks again, even though you should each have had a direct response from me by now.
Desiccant "packaging": Almost everyone suggested that I put my molecular sieve inside dialysis tubing to keep the dust out of the ethanol. Well, I've tried that in the past but the dialysis tubing we get here is squashed flat and *very* hard to open into a tube in its dry state. However I've now been told to tie off one end of the flat tubing then blow compressed air or similar into the open end. Thanks for that one, Joan.
What desiccant to use: Mostly respondents recommended molecular sieve, either alone or with silica gel as an indicator. There is also at least one proprietary brand of mol. sieve with a built-in indicator (probably not readily available in New Zealand).
Mel suggested dried CuSO4 which is self-indicating, turning from white to blue when it absorbs water.
One suggestion was to put magnesium metal in the ethanol to react with the water. I wasn't sure about that one - after reading the Merck Index I thought that if enough water was present the result *could* be a solution of Mg hydroxide with an alkaline pH.
Alternatives to solid desiccants: Bo and Tobias mentioned acidified DMP (2,2-dimethoxypropane), either as an additive to the ethanol or as a stand-alone dehydrating agent. Apparently acid.-DMP reacts with water to form acetone and methanol. Thanks guys, it is probably good as an additive but I want to keep things cheap and easy! The little I've read about DMP hasn't recommended it as a dehydrating agent on its own (extracts lipids, I think), and again there is the problem of increased expense.
Mark suggested fitting a plunger dispenser to the bottle of ethanol as soon as it is opened, combined with molecular sieve. Stewart suggested something similar but with calcium chloride in the air intake instead of mol. sieve in the ethanol.
What I have decided to do: Well, pretty much nothing actually. For reasons of economy and convenience I preferred the suggestions of either mol. sieve or dry CuSO4 in dialysis tubing, but I then asked a professor of chemistry which he would choose and he wasn't enthusiastic about either of them! He thought molecular sieve *might* change the pH "though it shouldn't" and that CuSO4 "probably isn't very efficient".
So whenever I open a new winchester (2.5 litres) of ethanol I will simply decant it into smaller (500ml/1 pint) oven-dried bottles, and then open them one at a time to top up the 125ml bottle that is part of the dilution series in the fume hood. For others in the dilution series (up to 95%) we will continue to use commercial "drum" ethanol.
So that was a surprise ending to this story, wasn't it?
Postscript: It turns out that this is exactly what The Southernmost EM Unit In The World (in Dunedin, New Zealand) has been doing with their ethanol for some time. Thanks for your message Allan, and I love the new signature file.... is that a yellow-eyed penguin? - I only have a mono screen :-) .
Regards
Stephen Edgar
Electron Microscope Unit, Pathology Department School of Medicine University of Auckland Private Bag 92019 Auckland New Zealand
A friend told me many people working on Spectroscopy and Microscopy are interested in mini-laser (cw, 532 nm and 1064 nm) and optical parametric oscillator (OPO, pulsed, tunable from 210 nm to 2500 nm). Now, we are making mini-laser, OPO and optics for laser users. Can anyone tell me the parameters of OPO and mini-lasers needed in Spectroscopy and Microscopy?
Thank you in advance for your help.
Jiwu Ling CASIX, Inc., PO Box 1103, Fuzhou, Fujian 350014, China Fax: 86-591-366-6957 E-mail: casix-at-public.sta.net.cn casixus-at-aol.com
Answer, Sample prep - polymer nanoparticles Latex particles have a very low contrast in TEM. They may also be filmforming. Therefore the best way to look at them is to use negative staining. The latex should be diluted until it is almost clear. A water solution of Uranyl Acetate (0.5%) is added to the latex. Place a small drop on a formvar coated grid (400 mesh). When you look in the TEM you should see dark rings around the particles. You may have to try a few times until you obtain the correct dilution. Good Luck! Helen Hassander
On Nov. 7, Alfred Kracher asked for sources of light-element microprobe standards, like nitrides and borides of heavy metals. I have come up with the following: Boron Nitride Lanthanum Hexaboride Titanium Nitride Silicon Nitride Boron Trioxide Boron Carbide Please e-mail me directly for boron and nitride percentages in these compounds. Steven Slap, Vice-President Energy Beam Sciences
Answer, Sample prep - polymer nanoparticles Latex particles have a very low contrast in TEM. They may also be filmforming. Therefore the best way to look at them is to use negative staining. The latex should be diluted until it is almost clear. A water solution of Uranyl Acetate (0.5%) is added to the latex. Place a small drop on a formvar coated grid (400 mesh). When you look in the TEM you should see dark rings around the particles. You may have to try a few times until you obtain the correct dilution. Good Luck! Helen Hassander
At 04:45 PM 11/9/95 EST-10, you wrote: } } ....Info from our virus experts is that it is not possible to spread by } email unless there is an attached file which is actually run.....
San Joaquin Delta College in Stockton, CA offers a 2 yr program certification in microscopy. We will be adding training in focused ion beam next semester as well. We have done so since 1970 and have graduates all over the United States. If you send me your address, I will send info. Sincerely, Judy Murphy
Judy Murphy, PhD Dept. of Microscopy San Joaquin Delta College 5151 Pacific Ave Stockton, CA 95207
To MSA subscribers I am an undergraduate student seeking higher education in the fields of SEM and TEM. I will be graduating soon and I am searching for a college that will offer certification in EM. Can anyone help me in this area?
} To MSA subscribers I am an undergraduate student seeking higher } education in the fields of SEM and TEM. I will be graduating soon } and I am searching for a college that will offer certification in EM. } Can anyone help me in this area? }
Everyone knows that the only place to go for EM training is San Joaquin Delta College in Stockton, CA. They have a fully accredited two year course for SEM and TEM of biological, or crystalline, materials microscopy. The address for the school is: San Joaquin Delta College Electron Microscopy Lab Holt 121 5151 Pacific Ave. Stockton, CA. 95207 Tel: (209) 474-5246
The head of the crystalline materials program is Dr. Frank Villalovoz and the head of the biological materials program is Dr. Judy Murphy. If you need more information ask around because we graduates of the program are everywhere.
Good Luck.
Michael W. Odum Spec. Prep. Tech. Gatan, Inc. 6678 Owens Dr. Pleasanton, CA. 94588 Tel: 510-463-0200 Fax: 510-463-0204 E-Mail: modum -at-gatan.com
We are having to get rid of our darkroom to make way for another instrument THUS we need to replace the functions of the darkroom. I would appreciate information about processors that can do both paper (RC papers)and film (EM film 4489, SO163, Kodak 4127 and Ektapan). I am familiar with the Mohr processor but need to know if there are any other good ones out there. Thank You in advance for the info, Judy Murphy
Judy Murphy, PhD Dept. of Microscopy San Joaquin Delta College 5151 Pacific Ave Stockton, CA 95207
With a problem of this kind you can not narrowly focus your TEM beam but you must spread it out over a very large area--- This way you reduce local thermal gradients nearyour intended picture location! Plastics films are always difficult because they heat and expand in the electron beam and change their height and focus!
George C. Ruben Dept Biological Sciences Datmouth College Hanover, NH 03755 tel 603 646-2144 fax 603 646-1347 ---------------------------------------- Dear Microscopists,
I'm looking at serial sections of nerves on formvar (1% in chloroform) coated slot copper grids (pretty standard stuff). The grids are acetone and distilled water cleaned several times and coated as per standard (which has worked consistently well for 4 years). For the past few months I've been having considerable trouble with what appeared to be focus problems. It now appears that the film is "shifting". The film is carbon stabilised and clean; we've tried modifying formvar coating preparations, carbon coating and specimen storage, but the film movement is still present. It appears "tight" when on the grid, but appears to shift under the beam (as indicated by what appeared to be focussing problems). Anyone with a suggestion????????????????????????/
} Dear Microscopists, } } I'm looking at serial sections of nerves on formvar (1% in chloroform) } coated slot copper grids (pretty standard stuff). The grids are acetone and } distilled water cleaned several times and coated as per standard (which has } worked consistently well for 4 years). For the past few months I've been } having considerable trouble with what appeared to be focus problems. It now } appears that the film is "shifting". The film is carbon stabilised and } clean; we've tried modifying formvar coating preparations, carbon coating } and specimen storage, but the film movement is still present. It appears } "tight" when on the grid, but appears to shift under the beam (as indicated } by what appeared to be focussing problems). Anyone with a } suggestion????????????????????????/ } } Thanks, } ______________________________________________ } } Shaun Sandow, } Division of Neuroscience, } John Curtin School of Medical Research, } Australian National University, } ACT 0200 } } Ph. (06) 249 4782 } Fax. (06) 249 2687 } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } You might try "Grid Glue"
The recipe can be found under "Tips & Tricks" at http://www.biotech.ufl.edu/~emcl
Just click the Wizard -at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at- Greg Erdos Phone: 904-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax 904-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 -at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at-
I would have to agree with Mary, and say don't give up the Polaroid yet, but with the following considerations (Which, since I'm not intimately familar with the LEO SEM, you should look at):
(1) SEM's with direct analog output to the HR-photo screens, and the
polaroid NEGITIVES are generally capable of 2500 lines of resolution,
at typical SEM photo-rates. 2k x 2k is still alittle less than that but you generally never use all 2500 lines on a neg. anyway.
(2) generally printers (1200dpi laser, or 300/600dpi dye-sublimation) can not ultimately give the same resolution as the polaroids.
(3) On some digital SEM's the output to the HR-photo screen is not the original analog signal, but rather the digitally captured image at its resolution (1k x 1k or 2k x 2k, more?). Therefore, if you send a 1k x 1k digital image to a polaroid HR-photo CRT capable of 2500 lines, you will not be getting the full resolution of the polaroid anyway so you might want to leave off the polaroid.
(4) Polaroid negs are very useful for a variety of photographic imaging needs, which can not be met with affordable digital output devices. It would be very useful to not only output current specimen images (i.e. specimens in the scope) but also stored images from disk to the HR-photo CRT (even images not generated on the scope? or after manipulation?) as if the Photo CRT is another digital output device like a printer. For this a 35mm HR digital output device might serve equally as well, and more usefully for other applications (i.e. slides for presentations?), however as was just discussed on the list a few weeks ago these can be very pricey ($6-10k), but might be an option instead of the polaroid [35mm + good grey scale + dye sub printer = polaroid output ? $ ?]
Pat Masarachia writes about problems when labeling sections through LR White embeded block of tissues and cells. She assumes that either the primary antibodies, which seem to have been properly stored, or the further polymerization of the block are at fault. However, there is no description of the visualization probe that is being used. If the probe is a gold-conjugated immunoglobulin then this may be the explanation for there being no EM labeling. This is because these gold probes are very unstable and the protein will separate from the gold over time (this starts even before purchase). One result of this is that the labeling efficiency (or sensitivity) of the probe is gradually reduced over time. It is possible also for the whole probe to just stop working. If the labeling protocol has stopped working for EM and LM I would suspect the latter.
Personally, I think that the best strategy for routine immunolabeling is to buy only protein A-gold (5 and 10 nm particle sizes) and use only these two probes for all labeling protocols. Only by doing this can the probes be constantly monitored for activity. If immunoglobulin-gold is used, firstly a service lab has to buy enough probes to cover all potential experiments (anti-rabbit, anti-mouse, anti-rat, anti-guinea pig, 5 nm and 10 nm particle size) and secondly, each probe must be tested for activity if it has been stored for long periods.
I just threw that last part in to add a little controversy and perhaps start a debate about the pros and cons of different gold probes and perhaps what are the best storage conditions for primary antibodies and colloidal gold probes. I know there are exceptions to using protein A-gold and some advantages to using gold-conjugated antibodies but most of us seem to be placed in a role where we have to help others at least some of the time. Under these conditions simple is best.
Sorry Pat for the long reply. Feel free to contact me if you still have questions.
Paul Webster Center for Cell Imaging Yale School of Medicine http://info.med.yale.edu/cellimg Tel: (203) 785 3219
We are looking for a used mounting press for making metallographic samples from bakelite. We won't be running a large volume so a manual press is fine. As long as it is working we'll consider it. Please let me know if anyone has one that they'd like us to take off their hands!
Thanks!
David Henriks TEL: 800-728-2233 (toll-free in USA) South Bay Technology, Inc. 714-492-2600 1120 Via Callejon FAX: 714-492-1499 San Clemente, CA 92673 USA e-mail: 73531.1344-at-compuserve.com
} ... For the past few months I've been } having considerable trouble with what appeared to be focus problems. It now } appears that the film is "shifting". The film is carbon stabilised and } clean; we've tried modifying formvar coating preparations, carbon coating } and specimen storage, but the film movement is still present. It appears } "tight" when on the grid, but appears to shift under the beam (as indicated } by what appeared to be focussing problems). Anyone with a } suggestion????????????????????????/ } I have had good luck with specimens which shift under normal beam conditions by using very low beam currents (~10 pA). In order for this to give you a decent image, you need either to have a long exposure time or to use very sensitive film. LoDose or MRF32 will be sensitive enough, but they have large grain--there is still no free lunch. If you have a very sensitive video system, that will make focusing easier--we have an intensified CCD which gives a useful image at video rates, so focusing can be done in a few seconds. I suppose that using even a large mesh grid would not be satisfactory, but if it is, then that may also stabilize your specimen. Good luck. Yours, Bill Tivol
Just a reminder that unless you really need to be certified, there are good schools that teach SEM technique such as New Paltz and McCrne Research Institute in Chicago.
--------------------------------------
} To MSA subscribers I am an undergraduate student seeking higher } education in the fields of SEM and TEM. I will be graduating soon } and I am searching for a college that will offer certification in EM. } Can anyone help me in this area? }
Everyone knows that the only place to go for EM training is San Joaquin Delta College in Stockton, CA. They have a fully accredited two year course for SEM and TEM of biological, or crystalline, materials microscopy. The address for the school is: San Joaquin Delta College Electron Microscopy Lab Holt 121 5151 Pacific Ave. Stockton, CA. 95207 Tel: (209) 474-5246
The head of the crystalline materials program is Dr. Frank Villalovoz and the head of the biological materials program is Dr. Judy Murphy. If you need more information ask around because we graduates of the program are everywhere.
Good Luck.
Michael W. Odum Spec. Prep. Tech. Gatan, Inc. 6678 Owens Dr. Pleasanton, CA. 94588 Tel: 510-463-0200 Fax: 510-463-0204 E-Mail: modum -at-gatan.com
------------------ RFC822 Header Follows ------------------ Received: by qmgate_backup.anl.gov with SMTP;9 Nov 1995 12:55:17 -0600 Received: (from daemon-at-localhost) by Sparc5.Microscopy.Com (8.6.11/8.6.11) id LAA05983 for dist-Microscopy; Thu, 9 Nov 1995 11:15:08 -0600 Received: from core.gatan.com (core.gatan.com [204.119.73.10]) by Sparc5.Microscopy.Com (8.6.11/8.6.11) with ESMTP id LAA05979 for {Microscopy-at-sparc5.microscopy.com} ; Thu, 9 Nov 1995 11:15:06 -0600 Received: from [204.119.73.207] ([204.119.73.207]) by core.gatan.com (8.6.12/8.6.9) with SMTP id JAA08372 for {Microscopy-at-MSA.Microscopy.Com} ; Thu, 9 Nov 1995 09:10:41 -0800 Message-Id: {199511091710.JAA08372-at-core.gatan.com}
ways to get better fixing of films onto grids include:
don't use solvents to "degrease" grids. Many solvents leave a residue. Instead, whisk grids quickly through a small (ethanol burner, low bunsen) flame so they flash red hot. Too hot and you will know! This is guaranteed to make them clean and hydrophilic.
Put the film on the DULL side of the grid, which is smoother on the fine scale and offers better adhesion.
Use cellulose tape glue dissolved off in chloroform. (40 cm of tape in 100 ml solvent, must NOT be plastic tape, which softens & dissolves a bit) One drop dried onto each grid before applying the coating film.
Put on the carbon from a carbon arc at an angle to the grid, on the NON coated side, so the carbon forms a film connecting the bars to the film. Rotate the grids 180 degrees and put on another coat. That should anchor the film pretty firmly.
One thing to keep in mind when discussing analog (signal direct to record CRT) vs. digital (signal digitized first, then sent to CRT of a digital printer) recording methods is that the sampling process only limits the performance of the latter method. This is because, according to Nyquist, you must digitize a single line to 2000 pixels to reliably display data in which the smallest feature is 1/1000 th of the width of the image in size. (Not 1/2000!! which would require 4000 pixels/line).
It is true that, in either case, the image is pixellized in the vertical direction but pixellation makes the size of the smallest displayable element (i.e. its area in relation to the area of the whole image) about 3 times smaller in a 2500x2500 analog display than it is in a 2000x2000 digital display.
The difference will be most noticeable when recording images at very low magnification ( {2-5,000, depending on kV and spot size) where the resolution of the image is not limited by the size of the probing beam.
You should also keep in mind that printing out even a 2000x2000 image on a 300DPI dye-sublimation printer will require a print that is at least 6.6 x 6.6 inches on rather thick paper (which is usually 8.5 x 11 inches), a size that requires much more storage space than does a Polaroid print (and its negative, if you don't copy them onto 35mm negs as we do).
Oh, I know that "all storage will be done on disk", but how many big disc drives can you find now that will play data recorded even 5 years ago...(assuming that the media is still useable)?
Jim Pawley
} Skipping the HR-Polaroid output. } } I would have to agree with Mary, and say don't give up the Polaroid } yet, but with the following considerations (Which, since I'm not } intimately familar with the LEO SEM, you should look at): } } (1) SEM's with direct analog output to the HR-photo screens, and the } } polaroid NEGITIVES are generally capable of 2500 lines of resolution, } } at typical SEM photo-rates. 2k x 2k is still alittle less than that } but you generally never use all 2500 lines on a neg. anyway. } } (2) generally printers (1200dpi laser, or 300/600dpi } dye-sublimation) can not ultimately give the same resolution as the } polaroids. } } (3) On some digital SEM's the output to the HR-photo screen is not } the original analog signal, but rather the digitally captured image } at its resolution (1k x 1k or 2k x 2k, more?). Therefore, if you } send a 1k x 1k digital image to a polaroid HR-photo CRT capable of } 2500 lines, you will not be getting the full resolution of the } polaroid anyway so you might want to leave off the polaroid. } } (4) Polaroid negs are very useful for a variety of photographic } imaging needs, which can not be met with affordable digital output } devices. It would be very useful to not only output current specimen } images (i.e. specimens in the scope) but also stored images from disk } to the HR-photo CRT (even images not generated on the scope? or } after manipulation?) as if the Photo CRT is another digital output } device like a printer. } For this a 35mm HR digital output device might serve equally as } well, and more usefully for other applications (i.e. slides for } presentations?), however as was just discussed on the list a few } weeks ago these can be very pricey ($6-10k), but might be an option } instead of the polaroid [35mm + good grey scale + dye sub printer = } polaroid output ? $ ?]
**************************************** Prof. James B. Pawley, Ph. 608-263-3147 Room 1235, Engineering Research Building, FAX 608-265-5315 1500 Johnson Dr., Madison, WI, 53706 JBPAWLEY-at-FACSTAFF.WISC.EDU
-- [ From: Charles A. Garber, Ph. D. * EMC.Ver #2.10P ] --
On Nov. 7, Owen P. Mills wrote inquiring about the preparation of polymer particles in the range of 100-200 nm.
We do a large volume of these kinds of samples in our laboratory:
1] If they are clumping, then probably the concentration of the suspension is too high and you need a higher dilution by at least one and maybe two or three orders of magnitude.
2] You want to use an "aerosol" type "duster" valve that incorporates a capillary tube to disperse the now highly diluted suspension. The best (but certainly not the only) one I know of is made by Ernest F. Fullam, Inc. in Schenectady, NY. The cost is less than $100. This is in all probability the "aerosol" method you mentioned in your posting.
3] You have to worry about the glass transition temperature. If above room temperature, then you need not further worry. If below room temperature, then the particles at room temperature will be soft and you have to worry about the need to "harden" them up. Acrylic particles can be "hardened" for example by UV using quartz irradiation tubes.
4] While particles in this range could in theory be done by SEM, especially cryo SEM, you will find that Pt/C replication TEM will give much more acceptable results. Use a shadowing angle of 45 degrees. You can measure the "shadow" and also the apparent diameter, and then compare the measurements as a insight into any "collapse" of the particles, being sort of a built in validation that the particles are indeed hardened into "hard" rigid marbles.
You do not need anything fancy in the way of a vacuum evaporator, any system giving a vacuum down in the "mid to low 10 to the minus five" range will give acceptable results.
5] If graft polymer latex, then if you want to see the morphological details of the graft vs. substrate polymer phase, you will have to use a still different technique to bring out such structures. But again, the method actually used will be determined by the specific polymers present and the kind of reactivity they exhibit.
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: GVKM07A-at-prodigy.com West Chester, PA 19381-0656 USA Customer Service: SpiSupp-at-aol.com
Does anyone know of a source where I can obtain a replacement bulb for a darkroom sodium vapor lamp? I found one in the Ted Pella catalogue, but $103 seems a little steep.
An error was made earlier in response to Shaun's question about slot grid problems. We use 0.3% (not 3%) formvar in chloroform on beryllium-copper grids. Sorry.
Paul webster raised the issue of the proper way to store gold probes. At least two EM supply companies (one being us) sell colloidal gold probes that have glycerol in the solution. These probes can be aliquoted and stored indefinitely in the freezer. More information on this is available at our WWW site: http://www.mwrn.com/ebs/ebs.com Steven Slap, Vice-President
} Does anyone know of a source where I can obtain a replacement } bulb for a darkroom sodium vapor lamp? I found one in the } Ted Pella catalogue, but $103 seems a little steep. } } Thank you, } } Dennis Shubitowski } dshubito-at-umich.edu
We get replacement bulbs for our Thomas safelights from the photographic supplier who has a purchasing contract with our university. They cost us $90, so $103 from Pella doesn't look so bad to me.
Judy Murphy asked: "I would appreciate information about processors that can do both paper (RC papers) and film (EM film 4489, SO163, Kodak 4127 and Ektapan). I am familiar with the Mohr processor but need to know if there are any other good ones out there".
Energy Beam Sciences manufactures an automatic EM film processor that works quite differently from the Mohr instrument. Somef you may have seen in our booth at the MSA meeting in Kansas City. For more information, see our WWW page: http://www.mwrn.com/ebs/ebs.com or contact me directly. Steven Slap, Vice-President
Hello Microscopists, I'd like to get your opinions on making a choice between a confocal system that can do triple label (green,red and high red emissions) or a deconvolution system. I realize this may be like comparing apples to oranges (the latter does not remove out of focus info but utilizes it), and demos will be conducted for both. So, if you had a wide range of uses (cultures, sections, yeast etc.) and had to choose one system, what would it be? We would be interested in both volumetric 3-D and thin optical sections. All comments are welcome.
Mediated negotiations have settled the strike at the University of Manitoba in Winnipeg, Canada, just in time to "save" the fall semester. Under the new agreement, individual professors cannot be targeted, and the faculty will determine which programs (not individuals, as proffered by the administration), will be eliminated in proven financial exigencies. Affected individuals would be offered options for redeployment, retraining, reduced appointment, or early retirement incentives. This strike of 1000 professors and professional librarians was a battle over the protection of academic freedom: from the start we offered cash concessions exceeding the budget shortfall.
We would like to thank you for the hundreds of worldwide e-mail letters, many reproduced in {http://www.xpressnet.com/umfa} . Internet made a critical difference in bringing international pressure on our administration and provincial government (which sided publicly with the administration), and we are grateful for your support. The issue of academic freedom was not discussed by the administration until we were 2 weeks into the strike, and your letters arrived. Our students came to realize that if academic freedom were compromised, their UM degrees would be worth less. We hope we have all now earned your respect after 3.5 weeks of collegial picketing, with supporters who flew in from universities across Canada, and with our students and local unions, at temperatures down to -17C in blowing snow. We hope we have set an example for the defense of academic freedom worldwide.
Please pass this on to your colleagues as appropriate. Comments may be sent to us at umfa-at-xpressnet.com, to President Arnold_Naimark-at-UManitoba.ca, and to Premier Gary Filmon: premier-at-leg.gov.mb.ca UMFA = University of Manitoba Faculty Association
We've been using off-the-shelf Low pressure lamps in our darkrooms for the last several safe lamps that have needed replacement. The last one was a Philips SOX 35 (35 Watts). Cost about $45.00 at our local industrial supply house. Hasn't fogged anything yet.
We are in the market for a new energy dispersive x-ray analysis system to attach to a Hitachi S520 SEM. The systems we are considering include:
LINK ISIS NORAN 3050 EDAX DX4 IRIDIUM II microanalysis system.
If any current users of these systems have any comments about the systems, we would most appreciate hearing from you.
Thanks in advance Dr Fiona Graham EM Unit University of Natal, Durban South Africa email: graham-at-ph.und.ac.za tel: +27 31 2602174 fax: +27 31 2616550
I have had a request from potential users to compile a list of vendors that provide software and hardware for time lapse video microscopy. I would appreciate any help in identifying and locating current vendors and products, and will gladly share the listing - please email me directly if you would like a copy.
TIA. Marc
Marc M. Friedman, Ph.D. Marketing Manager Olympus America Inc. Precision Instrument Division 2 Corporate Center Drive Melville, NY 11747-3157 Tel: (516) 844-5039 Fax: (516) 844-5111 email: marcfriedman-at-delphi.com
Hey folks -- My two cents on the question of gold labeled probes: We have had excellent results with Jackson (I have no connection with them) IgG gold (anti- rabbit, mouse, or guinea pig) which seems to be stable for at least a year at 4C. Since most of the labeling we do eventually involves localizing two antigens simultaneously, using IgG secondaries fits the "easiest is best" catagory.
Greg Martin Dept. of Cell Biology and Anatomy Johns Hopkins School of Medicine
} ... For the past few months I've been having considerable trouble with } what appeared to be focus problems. It now appears that the film is } "shifting". The film is carbon stabilised and clean; we've tried } modifying formvar coating preparations, carbon coating and specimen } storage, but the film movement is still present. It appears "tight" } when on the grid, but appears to shift under the beam (as indicated by } what appeared to be focussing problems). Anyone with a } suggestion????????????????????????/
Shaun, Is the movement only seen on slot grids or do you see on small mesh (200) also? The carbon stabilization should have done the trick and I am just wondering if the holder itself is unstable in some way. If focusing is the issue and not lateral movement, one VERY long shot could be the stability of the objective lens current. Sweeping the potentiometer knob repeatedly through the focus range can help clear dirt from the contacts and a shot of electrical contact cleaner behind the panel should be attempted before moving to more costly diagnostics. After all other low-cost options are exhausted, call for service.
I'm looking for tissue baskets that were/are used in the Polaron Critical Point Dryer. Ted Pella use to sell them (page 50 of their old catalog) as item # 1215-2, Tissue baskets only; set of 3. We use the baskets in our plunge freezing set-up and after a period of time they sprout legs and walk away from the lab...never to return! Ted quit selling them. Any suggestions? I will try to provide a good home for any old tissue baskets you may have stuffed away in a drawer and no longer need.
Best regards,
Beth Richardson EM Lab Coordinator Botany Department University of Georgia Athens, GA 30602-7271
I'm looking for tissue baskets that were/are used in the Polaron Critical Point Dryer. Ted Pella use to sell them (page 50 of their old catalog) as item # 1215-2, Tissue baskets only; set of 3. We use the baskets in our plunge freezing set-up and after a period of time they sprout legs and walk away from the lab...never to return! Ted quit selling them. Any suggestions? I will try to provide a good home for any old tissue baskets you may have stuffed away in a drawer and no longer need.
Best regards,
Beth Richardson EM Lab Coordinator Botany Department University of Georgia Athens, GA 30602-7271
As Dr Charles A Garber wrote there are special problems to image polymers in TEM. If you are not a polymer scientist, the negative staining method is the most safe one. It will give a proper result even if you have a film forming (low glass transition) or beam sensitive polymer. When you have learnt to dilute the latexes and add appropriate amount of Uranyl Acetate or PTA, it is also very easy to use. Helen Hassander Polymer group, Lund University
Greetings, How long are folks out there running their 50 Watt Mercury arc lamps? I ask because we used to run the 100 W arc's for 400 to 500 h, which is double or more than the manufacturer's guarantee of 200 h. The 50 W arc lamps are only guaranteed for 100h, which comes around awefully quickly. Any comments??
-- [ From: Charles A. Garber, Ph. D. * EMC.Ver #2.10P ] --
On Nov. 10, Beth Richardson asked about the CPD baskets once made by Polaron (e.g. their #1215-2). The last I heard, this product was available from Energy Beam Sciences, the US distributor for Fisons Instruments VG Microtech based in the UK.
As an alternative, SPI has made for some years our microporous specimen capsules in pore sizes of either 120um or 78 um. So far as I know, they have been used quite successfully in your application and with the advantage of there being far less of a chance of sample cross- contamination. See our web site (below) for further information under "Current WWW Specials". If still in doubt, e-mail me for further information and a sample pack for evaluation.
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: GVKM07A-at-prodigy.com West Chester, PA 19381-0656 USA Customer Service: SpiSupp-at-aol.com
--Boundary (ID qMMdyj3hoMjMtH004Fc9Rg) Content-type: TEXT/PLAIN
Thanks to everyone who responded to my call for suggested ways to prepare thin metal X-sections by electropolishing.
Naturally I assumed that EVERYONE would know which alloy system I'm working with, so I didn't include that bit of information. I'll let you in on the secret now though, we're talking aluminum.
To recap the responses:
Microtomy was suggested, but is unfortunately not an option due to the deformation it induces.
I was urged not to give up on tripod polishing (and I won't), but the boss wants to do away with the whole ion milling thing for this aplication (if possible).
A couple of individuals with either a better memory than mine, or a better database, not only remembered the combo Epol / Dimple experiment, but gave me a reference to check out.
Encapsulating in low temp melting metals was suggested. I've had some bad experiences with Pb / Bi in the TEM, so I'm a bit leary, but it's worth a look.
And the likely winner, to my mind, electrolytic or electroless coating.
I've actually had some good success in the past with electroless coating Ni on Al powders and ceramic particles, followed by ion milling. It wasn't much fun though (tended to coat everything but the bits of interest), and was awfully slow (maybe just my setup). I've yet to try Epolishing it either.
** The questions of the day then, to those of you in the know, is:
1. Can electrolytic coating be carried out as fast as electroless (and how)?
2. What metals would you suggest based on either speed or best chance to polish in the same electrolyte as Aluminum? Is Ni the best option?
Thanks in advance,
############################################################### # # # Don Steele STEELE-at-KRDC.INT.ALCAN.CA # # ALCAN INTERNATIONAL # # Kingston Research and Development Center # # P. O. Box 8400 # # Kingston, Ontario Canada K7L 5L9 # # # ###############################################################
Does Anyone know where I can get a copy of the "Monographs in Practical Electron Microscopy in Materials Science" parts 1-4, by J. W. Edington?
They were origonally published by the Philips Technical Library/MacMillan, but I heard the series was out of print.
Any suggestions would be helpful.
Thanks,
-Kirk ________________________________________________ Kirk A. Rogers krogers-at-materials.ecn.purdue.edu 317-494-8751 office http://materials.ecn.purdue.edu/~krogers 317-494-1204 FAX Purdue University, School of Materials Engineering, 1289 MSEE building, W. Lafayette, IN 47907-1289
We've run our last two 50W HBO Hg lamps for 1) just under 300 h (a mistake! - changed when the fluorescence got too dim) and 2) just over 200 h. As long as they don't get turned on and off too frequently they seem OK. It gives our Leica rep kittens, though.
cheers, Rosemary
____________________________________________________________ Rosemary White __ / Department of Ecology _/ \__/ \ and Evolutionary Biology / \ Monash University / Australia \ Clayton, Victoria 3168 \ ____ / phone 61-3-9905 5670 \_/ \_*_/ fax 61-3-9905 5613 __ email r.g.white-at-sci.monash.edu.au \/
Beth Richardson asked: "I'm looking for tissue baskets that were/are used in the Polaron Critical Point Dryer. Ted Pella use to sell them (page 50 of their old catalog) as item # 1215-2." Energy Beam Sciences has been, for almost two years, the exclusive U.S. distributor for Polaron equipment (and the authorized service representative). The tissue baskets, along with other supplies (targets, etc) are available, usually from stock. Please contact me directly for details. Steven Slap, Vice-President
Does anybody out there know about antibodies against SV40 (simian virus 40). The antibody should recognize the virus, not the simian virus 40 T antigen (SV 40 Tag). The antibody is needed for immunocytochemistry on the EM-level. Any information about researchers or commercial suppliers is greatly appreciated.
Thanks
Andreas Brech Electron Microscopical Unit for Biological Sciences Department of Biology, University of Oslo. P.O.Box 1062 Blindern N-0316 Oslo 3 Norway Tel.: + 43-22 85 61 89 (work) + 43-22 43 83 23 (privat) Fax.: + 43-22 85 47 26 e-mail.: abrech-at-bio.uio.no
Does anyone out there know of a simulation package for generating EELS spectra? NIST's DTSA does this for EDXS and I was just wondering about the possibility of one existing for EELS.
Many thanks in advance.
Colin V. ############################################################################### # # # Colin J. Veitch C.Veitch-at-geel.dwt.csiro.au # # Instrumentation Scientist # # CSIRO Division of Wool Technology Tel. +61 (0) 52 275611 # # P.O. Box 21 Fax. +61 (0) 52 275657 # # BELMONT Vic 3216 # # Australia # # # # "We see the Universe the way it is because if it were different, we would # # not be here to observe it." # # # ###############################################################################
Posted-Date: Sun, 12 Nov 1995 18:20:13 -0300 (BST) Received-Date: Sun, 12 Nov 95 18:20:14 BST
I have used "mount-quick" and found good results with it. There are two types: Mount quick solvent base Mount quick water base I've used the first one, and it is carried by EMS (fax # 215-646-8931 US). Hope it helps Adriana Rodriguez CENA/USP/Brazil
On Wed, 8 Nov 1995, Rosemary White wrote:
} Dear all, } } While in Germany a few years ago, a colleague bought a xylene-based medium } called Melanol for making permanent (or at least semi-permanent) mounts. } She has just about run out but can't find a supplier to buy more. Is there } a new supplier of this mountant? If not, can anyone advise re. a suitable, } less toxic substitute? } } TIA, } Rosemary White } } } } ____________________________________________________________ } Rosemary White __ / } Department of Ecology _/ \__/ \ } and Evolutionary Biology / \ } Monash University / Australia \ } Clayton, Victoria 3168 \ ____ / } phone 61-3-9905 5670 \_/ \_*_/ } fax 61-3-9905 5613 __ } email r.g.white-at-sci.monash.edu.au \/ } }
I'm sorry to tell you that antibodies can 'go bad' when "stored properly". It has been our experience that different antibodies exihibit different storage stabilities. The type of storage (eg. frozen w/ glycerol, storage where exposed to light or storage in "frost free" freezers) can also make a big difference. Even if you have done everything right, some antibodies just don't seem to store well. When you tell me that other antibodies are working well with your techniques, it seems likely to me that your antibody has degraded. Sorry to be the bringer (or confirmer) of bad news.
Dan
On Wed, 8 Nov 1995, Pat Masarachia wrote:
} Date: Wed, 8 Nov 1995 16:11:54 EST } From: Pat Masarachia {pat_masarachia-at-Merck.Com} } To: Microscopy-at-Sparc5.Microscopy.Com } Subject: LR White immunolabeling-LM/EM } } There is some rule that when something is going well don't change } or stop anything.. for several months I had good success immuno- } gold labeling using a polyclonal antibody for an integrin antigen. I } could get specific labeling in .3 micron sections of bone embedded } in LR White (silver intensified gold) for LM or in thin LR White } sections for EM. The same specific labeling results occurred in a } dozen experiments; antigen absorption of the antibody and deletion } of antibody were negative controls. The antibody was specific and } dependable. I had to interrupt the project and did not return for } a year. Murphy's law takes over. With the same antibody, using } aliquots stored at -20 C as per manufacturers instructions as well } as aliquots stored at -70 C, I cannot get labeling on sections } cut from the original blocks. The antibody still works on pre- } embedded tissue culture. On top of this, the original manufacturer } went out of business. The antibody produced by a new company with } the original protocol does not work. I thought that maybe the LR } White blocks after a year further polymerized to reduce antigenicity. } I worked up newly embedded tissue, freshly cut sections, and still } got no label. I've tried different secondary antibodies and different } blocking and different buffers -- no label. A different antibody for a } different antigen does specifically label in sections from old } blocks or new. So... can an antibody 'die' even if stored properly? } Sorry for this long winded tale but with no substitute available } commercially for this particular antibody I'd love to know what } happened. } Thanks for any comments. } } Pat Masarachia } Bone Biology } Merck Research Laboratories } West Point, PA 19486 } } }
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%% % % % Daniel Possin Work: 206/ 543-7489 % % Dept. of Ophthalmology RJ-10 FAX: 206/ 543-4414 % % University of Washington Home: 206/ 778-1714 % % Seattle WA 98195 USA Email: oemlab-at-u.washington.edu % % % % "The chinese expression 'cheung meng ba sui, gong hey fat choy' is % % equilvalent to Vulcan expression 'live long and prosper'. It's a % % small universe and getting smaller everyday". % % % %%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
There have been several running commentaries regarding resolution of various recording methods. I thought that I would ask the various vendors and others in the know what are the product specified resolutions AND recording areas of the following:
standard film (e.g. SO 163) CCD video cam
I was particularly interested in product specifics since they tend to be conservative estimates. I am also interested in how people think that these resolutions should be compared.
Daniel L. Callahan Department of Mechanical Engg. and Materials Science Rice University dlc-at-owlnet.rice.edu
At least two EM supply companies (one being us) sell colloidal gold probes that have glycerol in the solution. These probes can be aliquoted and stored indefinitely in the freezer.
Hello Steven,
Just a question about the storage of your gold probes in glycerin.
Then..... do you actually run the procedure with glycerin in the probe, or is there a clean up procedure after thawing the probe?
Have you found the glycerin to interfere with anything?
Thanks, Lou Ann
*************************** Lou Ann Miller Microscopic Imaging Lab College of Vet. Medicine University of Illinois 2001 S Lincoln Ave Urbana,Illinois 61801 217-244-1566 lmiller-at-ux1.cso.uiuc.edu
Microscopy Home Page: http://www.cvm.uiuc.edu/MicImagLab/MicImagLab.html
Personal Home Page: http://www.cvm.uiuc.edu/HomePages/LouAnnMiller/Homepage.html
We've used regular off-the shelf Na lamps for just that reason. The Philips SOX 35 fits the Thomas safe light housing and looks nearly the same as the OE lamp. They are about $45.00 at our local industrial supply house. Hasn't fogged anything yet.
Cheers, John heckman-at-pilot.msu.edu
} } } Does anyone know of a source where I can obtain a replacement } } bulb for a darkroom sodium vapor lamp? I found one in the } } Ted Pella catalogue, but $103 seems a little steep. } } } } Thank you, } } } } Dennis Shubitowski } } dshubito-at-umich.edu } } We get replacement bulbs for our Thomas safelights from the photographic } supplier who has a purchasing contract with our university. They cost us } $90, so $103 from Pella doesn't look so bad to me. } } John } chandler-at-lamar.ColoState.EDU } } }
I agree that it is a good idea to store colloidal gold probes in 50% glycerol. They can be left at -20#161#C for long periods without much loss of activity. As far as I know there has been no quantitative study on the effects of long term storage under these conditions on labeling efficiency. Once cryoprotected, the probes could also be stored at -80#161#C or in liquid nitrogen. I know of one report where colloidal gold probes were successfully freeze dried and reconstituted without harm (Lucocq and Baschong, some time in the '80's. They used lectin-gold conjugates. Does anybody know what the effects of long exposure to glycerol at RT has on gold probes and antibodies? I ask because I send antibodies and gold probes by mail in 50% glycerol.
In {v01520d00accd2b1877c1-at-[128.206.15.200]} , Tobias Baskin wrote: } Greetings, } How long are folks out there running their 50 Watt Mercury arc } lamps? I ask because we used to run the 100 W arc's for 400 to 500 h, which } is double or more than the manufacturer's guarantee of 200 h. The 50 W arc } lamps are only guaranteed for 100h, which comes around awefully quickly. } Any comments?? } } Thanks, } Tobias Baskin } } - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -
I find that I can run the 50 watt mercury arcs for about 170-190 hours before I find that they either begin to flicker or the intensity of the staining that I am using them to detect seems to drop in intensity. I have also been told, probably by an arc lamp salesperson, that if you run them too long beyond their normal life, they can explode. I have never had that happen in the 10 or more years that I have been using them.
Benjamin Walcott Dept of Neurobiology and Behavior University at Stony Brook Stony Brook, NY 11794
-- [ From: Charles A. Garber, Ph. D. * EMC.Ver #2.10P ] --
There has been some discussion on the subject of "Freon" being used in the HT tank. Of course, technically, there is no such thing called "Freon" being made any longer by DuPont (they still make this class of products but have given them different names). And because of environmental concerns, I don't think there is any more of the R-12 being produced either, by anyone, and in order to ship what is remaining in inventory,one almost needs an act of Congress to legally ship the R-12 over international boundaries. It was the R-12 that was originally used in the HT tanks.
We would advise against using either R-12 or any so-called replacements from the little "duster" sized cans (even though our firm has been a major supplier of such products) and that if these fluorocarbon materials are going to be used, then they should be procured only in the larger size containers, such as the 25 Kg container (not offered by SPI Supplies) referred to in a previous posting. Here is the main reason:
The "duster" type cans use at least one and sometimes two sets of internal "O" rings to seal the seams of the cans. These "O" rings, to give them the needed elastomeric properties are plasticized, and over time, the liquid inside leaches out some of this plasticizer. Many of us have done the "clean duster" test by turning the duster upsidedown and expelling against a clean glass surface and once the frost and water disappears, an easily observable film remains. It is this hydrocarbon film that could very well be causing the problem. And since there are no such "O" rings (so I am told) in the larger containers, that is why the problem seems to not show up when the larger container is used.
Apparently also, hydrocarbon contaminating oils can creep into the cans from the use of oil sealed compressors in the filling line. However, this source can be successfully controlled by having cans filled in lines for which this is never a problem. This also gives me the chance to point out that duster cans promoted, for example, for blowing emergency horns or chilling cocktain glasses, in general are not filled to the same standard of starting purity, and in general are not going to be the same quality product you would get from a reputable EM consumables supplier. Having said that however, the controlling factor is the leaching of plasticizer from the "O" rings.
I don't know about the moisture explanation, but it seems to be "accepted" that any level of hydrocarbon contaminant in the liquid in the HT tank is quite deleterious and that is why we have for some years advised against the use of these little cans for this purpose. Just how much plasticizer is going to be present is going to be a function of shelf life after filling, and it is also going to depend on the thermal history of the can during storage and shipment. However if someone has on their shelf any of the R-12 duster type cans, and is contemplating their use for this purpose, remember they were probably filled on the order of ten years ago (or more). My guess is that they would have for the HT tank application an unacceptable level of hydrocarbon contamination.
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: GVKM07A-at-prodigy.com West Chester, PA 19381-0656 USA Customer Service: SpiSupp-at-aol.com
Dear Mike D. This is a very timely question: deconvolution vs confocal. I have to make the same choice ... to some extent.
I have had a Scanalytics deconvolution demonstration and have also seen Deltavision. As you know, Scanalytics is based on the Fay and Carrington studies and Deltavision uses modified Aagard and Sedat mathematics. Both are good and one would have to decide which method you prefer. I have a Metamorph system (Universal Imaging) and found out that for a reasonable sum I can add the Scanalytics software to the system. You can use that after confocal imaging (of course defeating the usefulness of using many wavelengths and low intensity) but it may further sharpen your images. If you have a microscope with an xyz controlled stage, you can use the metamorph system to deconvolve (with Scanalyticson its. You now need a powerful PC, which could run overnight .... to deconvolve the images... and you will need a cooled CCD to collect the images.
I do believe there are times when the confocal microscope will be superior to the deconvolution and in reverse. I have a large multiuser facility in which to place the confocal and / or deconvolution equipment and have to have a system that is userfriendly and can give fairly quick results. I am fortunate in that I have a set-up on which I can test the Scanalytics for a fairly reasonable price. If it proves to be really useful, I will apply for funding for a faster computer, etc.
It is not an easy choice, but I believe if there is no confocal in the area, it is very useful and the best choice. What do others think?
There is an article in the latest issue of THe Microscope that might be of interest: "Cryo-Ultramicrotomy of Individual Latex Particles for Examination of Internal Morphology", Marcelli, Angela. The Microscope 43(3):117-120(1995)
Peter D. Barnett Forensic Science Associates e-mail: pbarnett-at-crl.com
Excuse me, but Delta is NOT the only school for EM training. I am a graduate of Madison Area Technical College and they offer a two-year Associate Degree program in electron microscopy. MATC grads are also everywhere!
Jean Ross Research Assistant Central Electron Microscopy Research Facility Univ. of Iowa Iowa City, IA 52242 319-335-8142
On Thu, 9 Nov 1995, Michael Odum wrote:
} On 11/7/95 Bridgett Byrnes sent: } } } To MSA subscribers I am an undergraduate student seeking higher } } education in the fields of SEM and TEM. I will be graduating soon } } and I am searching for a college that will offer certification in EM. } } Can anyone help me in this area? } } } } Everyone knows that the only place to go for EM training is San Joaquin } Delta College in Stockton, CA. They have a fully accredited two year course } for SEM and TEM of biological, or crystalline, materials microscopy. The } address for the school is: } San Joaquin Delta College } Electron Microscopy Lab Holt 121 } 5151 Pacific Ave. } Stockton, CA. 95207 } Tel: (209) 474-5246 } } The head of the crystalline materials program is Dr. Frank Villalovoz } and the head of the biological materials program is Dr. Judy Murphy. If you } need more information ask around because we graduates of the program are } everywhere. } } Good Luck. } } Michael W. Odum } Spec. Prep. Tech. } Gatan, Inc. } 6678 Owens Dr. } Pleasanton, CA. 94588 } Tel: 510-463-0200 } Fax: 510-463-0204 } E-Mail: modum -at-gatan.com }
} How long are folks out there running their 50 Watt Mercury arc } lamps? I ask because we used to run the 100 W arc's for 400 to 500 h, which } is double or more than the manufacturer's guarantee of 200 h. The 50 W arc } lamps are only guaranteed for 100h, which comes around awefully quickly. } Any comments??
Dear Tobias, I had some experience with a 1 kW high-pressure Hg lamp which may be relevant to your question. It too was rated at 100 hrs, and the mfr told me the usual method of failure was explosive! I set up the lamp and ran it continuously for } 400 hrs, and it was still good when I turned it off. The biggest contribution to failure comes from thermal stress, so if the lamp is not turned on and off, but is stabile at a particular tempera- ture, it will last a long time. The lamps usually fail just after they are turned on for one time too many. This happened to me just as I reached for the air vent on the lamp housing. I was startled but uninjured, but the lamp took out the reflector, damaged a lens and put several dents in the housing. If you can arrange your work so that it can all get done in one stretch, you should not be afraid to try using the lamp for 400-500 hrs, but if you are using it in equipment which is turned on only when in use, change the lamp when the mfr says, unless you have a surplus of housings and a shortage of lamps. Good luck. Yours, Bill Tivol
1x2 mm slot grids are notoriously hard to completely stabilize even when carbon coated. If you are sure of your coating technique, I would begin viewing each new grid by spreading the electron beam just wider than the length of the grid slot and irradiate the whole thing for about 10-15 minutes prior to taking a closer look. This will strip the light elements out of the formvar film and "carborize" it making it more stabile. If this still is not enough I would consider making thicker films by increasing the concentration of your formvar solution. 1% Formvar in chloroform seems a little thin to me even with carbon coating. If you still have trouble, e-mail me again.
Dan
On Thu, 9 Nov 1995, Shaun Sandow wrote:
} Date: Thu, 09 Nov 1995 16:00:56 +1000 } From: Shaun Sandow {shaun.sandow-at-anu.edu.au} } To: microscopy-at-Sparc5.Microscopy.Com } Subject: slot grid problems } } Dear Microscopists, } } I'm looking at serial sections of nerves on formvar (1% in chloroform) } coated slot copper grids (pretty standard stuff). The grids are acetone and } distilled water cleaned several times and coated as per standard (which has } worked consistently well for 4 years). For the past few months I've been } having considerable trouble with what appeared to be focus problems. It now } appears that the film is "shifting". The film is carbon stabilised and } clean; we've tried modifying formvar coating preparations, carbon coating } and specimen storage, but the film movement is still present. It appears } "tight" when on the grid, but appears to shift under the beam (as indicated } by what appeared to be focussing problems). Anyone with a } suggestion????????????????????????/ } } Thanks, } ______________________________________________ } } Shaun Sandow, } Division of Neuroscience, } John Curtin School of Medical Research, } Australian National University, } ACT 0200 } } Ph. (06) 249 4782 } Fax. (06) 249 2687 } }
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%% % % % Daniel Possin Work: 206/ 543-7489 % % Dept. of Ophthalmology RJ-10 FAX: 206/ 543-4414 % % University of Washington Home: 206/ 778-1714 % % Seattle WA 98195 USA Email: oemlab-at-u.washington.edu % % % % "The chinese expression 'cheung meng ba sui, gong hey fat choy' is % % equilvalent to Vulcan expression 'live long and prosper'. It's a % % small universe and getting smaller everyday". % % % %%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
I can assure you that Nyquist does apply to the sampling of images. Usually Nyquist is set by the spatial frequencies transmitted by the optics but in any case, Nyquist defines the smallest spatial feature that will be reliably preserved in digital data.
One is apt to think that one will be able to record a small feature as long as it is larger than the size of a pixel. If the feature happens to be situated so that its centroid is in the center of a pixel you will indeed record it (supposing perfect digitizing , something else that doesn't always happen). However, in general, it will not be so conveniently situated (if the feature lands on the border line, you will end up recording a feature that is twice as wide as it should have been and only half as bright) and the oversampling factor of 2 (2.3 actually) substantially improves the odds (at least if the contrast varies with spatial freguency in a smooth and reasonable manner).
The SEM is an interesting case in that this last condition is often not met. Although at high mag, the resolution in most recorded images is much more likely to be limited by too high a beam voltage or too large an electron beam (so Nyquist is irrelevant), the immense magnification range of the SEM, coupled to the fact that the beam diameter need not change with magnification, means that this is not true at low mag. Given a slow enough scan time (to provide more data for a good S/N for more pixels) and a good enough record screen (many do not really have the 2500 lines that they claim but 10,000 x10,000 is possible and ideed was once offered by Zeiss), the image bandwidth can greatly exceed the Nyquist frequency of any reasonable digitizer (by 10-100x). For instance, it is quite possible for adjascent pixels to be black then white, something that does not happen with a signal having a bandwidth near the Nyquist frequency.
Fortunately, this is not a major problem because, if you need 100 sec to get 2nm data from area 1000 nm on a side (100kx), then you will require 10,000 sec to acquire such data from an area 10,000 nm on a side (10,000x) and 1,000,000 sec to do the same at 1,000x and so one at lower mags. The fact that no one wants to wait this long (let alone the stability and focusing problems) means that the fact that we really don't have any way to store or display such data in not a problem.
Jim Pawley
**************************************** Prof. James B. Pawley, Ph. 608-263-3147 Room 1235, Engineering Research Building, FAX 608-265-5315 1500 Johnson Dr., Madison, WI, 53706 JBPAWLEY-at-FACSTAFF.WISC.EDU
A student here at the dental school is doing a project a bit removed from our normal lines of research. He needs to find a source for Anti-rabbit Alkaline Phosphatase primary antibody (monoclonal) and/or Anti-rabbit osteocalcine Ab (monoclonal) if I am reading his writing correctly.
So far he has not been able to find a source from the catalogs we have. Does anyone know where we can git htese materials?
The Edington series is now distributed by Techbooks in Fairfax (Herndon?), VA. They have combined the four (five?) books into one volume which sells for about $55. The original (Philips) series has been out of print for some time now. I am not sure that the images in the Techbooks version are as good as in the original, but the books are still an excellent resource. Their phone number is (703) 352-0001.
Dick Fonda
krogers-at-ecn.purdue.edu (Kirk Rogers) wrote } } Does Anyone know where I can get a copy of the "Monographs in Practical } Electron Microscopy in Materials Science" parts 1-4, by J. W. Edington? } They were origonally published by the Philips Technical Library/MacMillan, } but I heard the series was out of print. } Any suggestions would be helpful. } } Thanks, } -Kirk } ________________________________________________ } Kirk A. Rogers } krogers-at-materials.ecn.purdue.edu } 317-494-8751 office http://materials.ecn.purdue.edu/~krogers } 317-494-1204 FAX } Purdue University, School of Materials Engineering, } 1289 MSEE building, W. Lafayette, IN 47907-1289
_____________________________________________________________ Richard W. Fonda Naval Research Laboratory (202) 767-2622 Code 6324 _____________________________________________________________
One of my colleagues has some fluorescent wholemount and sections of tissues that were dehydrated and cleared in methyl salicylate. Does anyone have a good suggestion for a permanent mounting medium? Thanks in advance.
Thomas E. Phillips, Ph.D. Associate Professor of Biological Sciences Director, Molecular Cytology Core Facility 3 Tucker Hall University of Missouri Columbia, MO 65211 (314)-882-4712 (voice) (314)-882-0123 (fax)
The price is $55. It is a fair quality single volume reproduction printed in India. Philips told me a number of years ago that they had lost the masters, so I think that this is about the best quality you can get.
Cheers, Henk
} } Does Anyone know where I can get a copy of the "Monographs in Practical } Electron Microscopy in Materials Science" parts 1-4, by J. W. Edington? } } They were origonally published by the Philips Technical Library/MacMillan, } but I heard the series was out of print. } } Any suggestions would be helpful. } } Thanks, } } } -Kirk } ________________________________________________ } Kirk A. Rogers } krogers-at-materials.ecn.purdue.edu } 317-494-8751 office http://materials.ecn.purdue.edu/~krogers } 317-494-1204 FAX } Purdue University, School of Materials Engineering, } 1289 MSEE building, W. Lafayette, IN 47907-1289
Henk Colijn colijn.1-at-osu.edu OSU Campus Electron Optics Facility ------------------------------------------------------------------- An optimist believes that we live in the best of all possible worlds. A pessimist fears this is true.
The price is $55. It is a fair quality single volume reproduction printed in India. Philips told me a number of years ago that they had lost the masters, so I think that this is about the best quality you can get.
Cheers, Henk
} } Does Anyone know where I can get a copy of the "Monographs in Practical } Electron Microscopy in Materials Science" parts 1-4, by J. W. Edington? } } They were origonally published by the Philips Technical Library/MacMillan, } but I heard the series was out of print. } } Any suggestions would be helpful. } } Thanks, } } } -Kirk } ________________________________________________ } Kirk A. Rogers } krogers-at-materials.ecn.purdue.edu } 317-494-8751 office http://materials.ecn.purdue.edu/~krogers } 317-494-1204 FAX } Purdue University, School of Materials Engineering, } 1289 MSEE building, W. Lafayette, IN 47907-1289
Henk Colijn colijn.1-at-osu.edu OSU Campus Electron Optics Facility ------------------------------------------------------------------- An optimist believes that we live in the best of all possible worlds. A pessimist fears this is true.
Hello to all, I was wondering if anyone out there has a used photometrics PXL cooled CCD (preferably with the kodak 1400 chip), that they would be willing to part with. Please contact me at the above address or call, and we can discuss terms. Thank you.
Michael Delannoy Microscopy Facility JHMI Baltimore, Md. (410) 955-1365
A lab in our Biochemistry department is interested in labelling a membrane protein, found in the inner membrane of e-coli. At the present time, I don't have a set-up for embedding in Lowicryl. So has anyone used LR white for thin section immunoAu labelling of proteins found in e-coli? Or, has anyone used Nanogold/ silver enhancement to label proteins in e-coli, before embedding. We are experts in TEM and a variety of labelling techniques, but novices with e-coli; so specific references or preparation methods would be greatly appreciated.
Louisa Howard EM Facility/ Remsen 240 Dartmouth Medical School Hanover, NH. 03755
Message-Id: {199511141857.NAA23038-at-thomas.ge.com} X-Authentication-Warning: thomas.ge.com: daemon owned process doing -bs
Ref.: Laser vs. digital deconvolution
I am not associated with the company that makes the product and gives this information as a user's advice.
You may want to check VayTek, inc. (514 472-2227) headed by John Kesterson, Ph.D. Their Micro-Tome deconvolution softwares costs 10-20K and are good. Thus, if you already have a good microscopic setup..... It is possible that VayTEk goes into partnership with ONCOR (1-800 77-ONCOR), but I am not sure on details. I will be interested on the outcome of the results, and perhaps you can compile relevant messages and make them available because I too may purchase a digital decovulating package, only thing I need to add confocal to my lab.
****************************************************************** *Cesar D. Fermin, Ph.D \|T|/ Fax (504) 587-7389 * *Tulane Medical School /|M|\ Voice Mail (504) 584-2618 * *Pathology/SL79 \|C|/ Secretary (504) 584-2436 * *New Orleans La 70112-2699 /|*|\ Lab/Techn. (504) 584-2521 * * {Fermin-at-tmc.tulane.edu} Dir. Morphological Services * ******************************************************************
Sender: kayton-at-ohsu.edu (Robert Kayton) Message-Version: 2 } To: microscopy-at-Sparc5.Microscopy.Com
for those who are intested in laser printing
the following company in New York has a very good alternative to print images on Laserjet 3/4/4+ printers - it is a German interface board with 150, 300 and even 600 dpi printing capability, with a palette of 256 from 1024 gray levels. it works really fine for printing hires SEM images and prints a 2 MByte image in seconds!
the experience shows that it is a real alternative to photographic images when you have to find another way to print good pictures from a SEM/STEM.
contact : Electro Image, Inc. 9, Round Hill Road LAKE SUCCESS, NEW YORK 11020 tel 516 773 4305 fax 516 773 2955
} One of my colleagues has some fluorescent wholemount and sections of } tissues that were dehydrated and cleared in methyl salicylate. Does anyone } have a good suggestion for a permanent mounting medium? Thanks in advance. } } } } Thomas E. Phillips, Ph.D.
I have successfully used methlysalicylate--the final clearing solution--for long-term storage of fish cleared this way. You might also try transferring them to 100% glycerin, probably through a graded series, but I have not tried that and don't know how well it would work.
&&& Illigitimi non carborundum &&&&&&&&&&&&&&&&&&&&&&&&&&&&&&&&& Philip Oshel Center for Electron Microscopy University of Illinois (217)244-3145 oshel-at-ux1.cso.uiuc.edu
Sender: kayton-at-ohsu.edu (Robert Kayton) Message-Version: 2 } To: microscopy-at-Sparc5.Microscopy.Com
Dear Fellow microscopists,
Probably the question I'm going to ask you has already been answered several times, but up to now we did not bother about it, so that I surely deleted all related mails that have probably been sent.
In our laboratory we are thinking about changing the procedure for printing our TEM negatives (although we do some minor work with the SEM). Up to now we use the standard printing on paper, but as the amount of pictures we are generating is always increasing, we think that we should change and start with the procedure of digitazing (the negatives) and printing them. We are thinking about purchasing a flat scanner to scan the negatives and a high resolution printer (perhaps an ink sublimation printer). We would appreciate receiving comments on this subject and to hear the experience of other laboratories: which scanners are OK or which should be the minimum resolution of them, which printer type and resolution, computer and physical support to keep the images (writeable) CD, ..., need for an image treatment software, aso.
Thank you in advance,
Albert Romano-Rodriguez
EME, Electronic Materials and Engineering Dept. Applied Physics and Electronics University of Barcelona Diagonal 645-647 E-08028 BARCELONA Spain e-mail: romano-at-iris1.fae.ub.es tel: +34 3 402 11 47 FAX: +34 3 402 11 48
Sender: kayton-at-ohsu.edu (Robert Kayton) Message-Version: 2 } To: microscopy-at-Sparc5.Microscopy.Com
In response to a number of questions received on the Lexmark Optra R: 1) Printing speed is 16 ppm at 600 dpi and 8 ppm at 1200 dpi and uses a 32-bit RISC processor. 2) To print a full 8.5" x 11" page of text, takes about 20 seconds from "go". 3) Toner comes in two packages - one for approx. 7000 pages is $179.95 and the other, for approx. 14000 pages is $265.95. These prices are out of the Misco catelog and one may be able to do better. 4) I previously mentioned the option of providing additional Printer Memory (I called it "RAM"). One can also purchase a Flash Memory Option to store information like downloaded fonts and macros. 5) The printer has two connections on the system board that can be used for either a Disk Option or a Network Option (or two Network Options, or one of each). Disk Option: A 40MB hard disk and a thumbscrew - for example, for downloaded fonts and macros. Network Option: To connect directly to a LAN - Token-Ring, Ethernet (10BASE2 or 10 BASE-T) or LocalTalk (whatever that is?). This option consists of a network card, a thumbscrew, a diskette containing the Network Printer Utility and documentation. You will need to provide the appropriate network cable. 6) I do not know the prices of any of the options but you can call (800)358-5835 for the name of a dealer near you. 7) In case of a problem, they will express an exchange printer OR repair and return yours. Apparently there is an on-site warranty repair service which I do not have. IBM, however, can provide local service. Lastly - in my previous note I may not have advised how REALLY easy this printer is to operate. I have had mine for a month now - and have not refered to the manual even once! Even in start-up, the printer "looked at" my computer software and adjusted itself to my previous printer driver. Prior to purchasing this unit, I looked at many high end laser printers and could not find any, at any price, that would even start to compare. I welcome any other inquiries, Don Grimes, Microscopy Today
Sender: kayton-at-ohsu.edu (Robert Kayton) Message-Version: 2 } To: MICROSCOPY-at-Sparc5.Microscopy.Com
1) Rapid (i.e.,in seconds) microwave fixation is used with success to fix plant and animal tissues. In addition, several reports show the benefits of using microwave fixation to preserve soft tissues encased by hard shells (e.g., clams, teeth, insects).
Recommended reading: Login, G. R., & Dvorak, A. M. (1994). Application of microwave fixation techniques in pathology to neuroscience studies: A review. J Neurosci Methods, 55, 173-182. Login, G. R., & Dvorak, A. M. (1994). Methods of microwave fixation for microscopy. A review of research and clinical applications: 1970-1992. Prog Histochem Cytochem, 27/4, 1-127. Kok, L. P., & Boon, M. E. (1992). Microwave Cookbook for Microscopists. (3rd ed.). Leyden: Coulomb Press.
2) Microwave curing of resins (acrylics and epon substitutes) is also successful. Recommended reading: Giammara, B. (1993). Microwave embedment for light and electron microscopy using Epoxy resins, LR White, and other polymers. Scanning, 15, 82-87. Login, G. R., & Dvorak, A. M. (1994). The Microwave Toolbook. A Practical Guide for Microscopists. Boston: Beth Israel Hospital.
3) Microwave processing is used by some large labs for rapid throughput of specimens. To date, no automatic microwave processors exist so- although microwave processing is rapid it is also a bit labor intensive. Recommended reading: Leong, A. S.-Y. (1991). Microwave fixation and rapid processing in a large throughput histopathology laboratory. Pathol, 23, 271-273. Leong, A. S.-Y. (1993). Microwave techniques for diagnostic laboratories. Scanning, 15, 88-98.
4) The major problems reported with microwave methods are lack of uniformity and reproduciblity. ALL microwave ovens have high and low energy areas. The literature reports several methods for calibrating microwave ovens to improve results.
Login, G. R., & Dvorak, A. M. (1994). The Microwave Toolbook. A Practical Guide for Microscopists. Boston: Beth Israel Hospital. Giberson, R. T., & Demaree, R. S., Jr (1995). Microwave fixation: understanding the variables to achieve rapid reproducible results. Microsc Res Tech, 32, 246-254. Gu, J., Forte, M., Hance, H., Carson, N., Xenachis, C., & Rufner, R. (1994). Microwave fixation, antigen retrieval and accelerated immunocytochemistry. Cell Vision, 1(1), 76-77.
5) Fortunately, many companies which sell Microscopy products have also designed, tested, and now sell microwave devices and accessories which improve microwave methods and most importantly make them safer to use in the laboratory.
Please contact me if you have additional questions.
Gary R. Login, D.M.D., D.M.Sc. Assistant Professor of Oral Pathology Beth Israel Hospital Department of Pathology 330 Brookline Avenue Boston, Massachusetts, 02215
I have previously worked for Zeiss for over 10 years in microscope sales - in that time I have found that the 100 DC HBO burners would run for 200 to 600 hours with the only problem of light output diminishing and problems then getting even illumination over the 175 to 225 hour mark. The 50W HBO AC is a different story - the buld is rated at 100 hours but depending upon operating conditions they will cloud up and darken as fast as 60-75 hours of use and I have had in 15 years in the indusrty 4 blow up violently - they do not go quitely like the 100 DC models. All four were in the 75 to 150 hour range. As such I have highly encouraged people to change at 100 hours!! - the Bulbs are same price but upon intial purchase the DC power suplly is considerably more expensive - you pay up front anf get long and safe bulb life or you save now and pay in the number of bulbs purchased. Depending upon manufacture the difference could be $300 for AC aupply vs. $2500 for DC supply. $2000 buys 10 to 15 bulbs and if you are a heavy fluoresence user than get the 100 if not the 50 is a good way to put more money into the optics where it belongs.
Scott E. Berman Advanced Imaging Concepts Princeton, NJ
Does anyone know of a supplier (Australian?) of carbon fibre for a Balzers model CED 010 carbon coating unit. The fibre supplied by Balzers has recently increased in price to A$95 for 3 metres. They have also changed their supplier and the new thinner thread gives a thinner coating in our system.
Thanks for any suggestions,
Dave
------------------------------------------------------------------ Dave Phelan Electron Microscope Unit Ph (049) 215667 [+61 49 215667] University of Newcastle Fax (049) 215669 [+61 49 215669] NSW 2308 Email emu-at-medicine.newcastle.edu.au AUSTRALIA ------------------------------------------------------------------
Does anyone know of a software which I can use to estimate the connectivity of phases of interest ? I have some SEM binary images showing pore distribution patterns, and would like to determine how well the pores connected to each other, based on their connectivity values.
Thanks.
Long Liang ARCO EPMA/SEM Lab Plano, TX lliang-at-is.arco.com
Hi, this is Lucy Yin from microscopy center at Amherst, Univ. of Mass. I would like to be on the listserver of MSA microscopy. Please tell me the procedure to get on . Thanks, Lucy 95-11-15
According to the 15 November edition of the Wall Street Journal (pg B8) the good times virus is no longer a hoax. The newest version of the e-mail has an attached file named either "install.exe" or "AOL Gold" which, when installed may render the hard drive inoperative according to an AOL spokeswoman. AOL plans on notifying all its members today.
-Kirk ________________________________________________ Kirk A. Rogers krogers-at-materials.ecn.purdue.edu 317-494-8751 office http://materials.ecn.purdue.edu/~krogers 317-494-1204 FAX Purdue University, School of Materials Engineering, 1289 MSEE building, W. Lafayette, IN 47907-1289
I'm looking for researchers using freeze substitution in their SEM and TEM protocols. I know there are several nice insturments for this protocol--I'ld like to contact researchers who are doing biological freeze substitution without the benefit of these expensive devices and who reside in the southwestern USA
I'ld appreciate any help in this regard
thanks in advance
steve barlow
--------------------------------------------------------------------------- ------------------------------------------------------------------------------ Dr. Steve Barlow EM Facility/Biology Dept. San Diego State University San Diego CA 92182-4614 phone: (619) 594-4523 fax: (619) 594-5676 email: sbarlow-at-sunstroke.sdsu.edu
Thank you for all the replies regarding replacement sources for the sodium vapor lamp bulb. National Graphic Supply did indeed carry them for $85 which seemed to be the best price around. I will also look into sodium lamps from home improvement stores which seems like a viable alternative.
Thanks again for all the suggestions!
Sincerely, Dennis Shubitowski University of Michigan School of Dentistry dshubito-at-umich.edu
At , you wrote: } Dear Fellow microscopists, } [ ... snip ... ]
} We are thinking about purchasing a flat scanner to scan } the negatives and a high resolution printer (perhaps an ink } sublimation printer).
} We would appreciate receiving comments on this subject and } to hear the experience of other laboratories: which scanners are } OK or which should be the minimum resolution of them, which printer } type and resolution, computer and physical support to keep the } images (writeable) CD, ..., need for an image treatment software, } also. } Thank you in advance, } } Albert, } We use the HP IIcx flatbed which is no longer available, but is now sold as a HP 3c ( {$1000). It is a very capable scanner, but you have to buy the transparency adapter for an extra $600. As a WIntel applet the twain software is very solid but they haven't delivered a 32bit driver yet for Win95/NT ... the 16bit still works however.
Regarding dye-sub printers ... I suppose you realize they are color printers and you pay $extra$ to print on color stock ( {$3/pg) in spite of the image being monochrome. The option does exist for grayscale ribbons ( {$2/pg), but they are more than a hassle if you use color sometimes and want to use grayscale at times. We ^do^ use a Codonics ethernet printer which has the Kodak dye-sub engine ... and we can reccommend it ... it is a different type of ethernet printer ... let me know if you want to know more about it. As another option there are Ag salt laser printers available which print grayscale at 256dpi which are ^not^ dithered ... ie, ^true^ 256 shades of gray. They are expensive ( {$14k), but the cost is less than a $1/pg.
Regarding, archival storage of image files ... you should $invest$ in magneto-optical. We chose to go with a Fujitsu 230MO drive ( {$700) for which the replacable media costs {$30 and can be found for near $20. The other option which we now wished we could have afforded would have been to go with 1.3Gb drives for which the drives are more expensive, but the media costs are less expensive (per Mb). These MO drives offer the freedom of write/delete ..., a third option would be write once CR-ROM drives for archiving ... again the drives are expensive but the media is less than $15 for 600Mb. Lastly, in this regard and to mention alternatives, eg Zip drives, some people would imply they shouldn't be used for "archiving" as the magnetic media shouldn't be trusted to any degree more than floppies or hard drives for occasional and intangible influences. On the other hand, MO drives require an annealing temperture to change a bit, which also means they write more slowly but they do read quickly.
Hope this helps ..
cheers, shaf
{\/} /\ {\/} /\ {\/} /\ {\/} cognito, 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/epmahome/
We have a Philips 201C TEM that we must get rid of to free-up some space in our lab. If you are interested, please contact me and we can discuss the details of its availability. Thanks! Donna Wagahoff SIU School of Medicine P.O. Box 19230 Springfield, Il. 62794-1220 phone 217-782-0898 fax 217-524-3227
**************************************************************** I am sending this message to the list as a whole since I mistakenly used that forbidden REPLY button and I think it went to only one recipient and I had hoped that it would be more generally informative. ****************************************************************
The question was about using arc lamps for more than the rated life. The characteristics of arc lamps are the key to understanding what may be reasonably done with them, but the wide variey of power supplies that are available are the other part of that equation and often little information is available to help users in this regard. Armed with the detailed characteristics of mercury arc lamps, please go after your manual or vendor for the missing part so that you may make the right decisions.
Mercury arc lamps are given a nominal lifetime rating based on 30 min of operation per start and operation at the specified conditions. Each ignition of a mercury arc does some damage to the electrodes. A sufficient current at a high voltage is required to break over the arc in the cold lamp where the mercury is not in the high pressure vapor state. This energy serves to vaporize some mercury and get the arc established, at which point it can be maintained at a lower current and voltage. This large surge erodes the tips of the electrodes, opening the spacing a little each time. Both in starting and in operation, the electrode spacing is involved in establishing the arc voltage at the operating current. When the spacing is too large, ignition and operation become more difficult as described below. The ignition circuitry that Osram, for example, describes in their literature of a few years back, is simple passive electronics that simply do the job. More modern electronic units CAN start the lamps more gently and this, together with longer operation per start, will be reflected in longer usable lamp life.
Mercury arc lamps have nominal ratings: a 100W lamp operated at the nominal 5 amps current will only give 100 watts at one brief part of its cycle. This is because of the change of the arc electrode spacing (and thus arc voltage). Most early power supplies, whether DC or AC used a choke to limit and stabilize the lamp current; since the arc voltage is much lower than the line voltage, this gave an approximation of constant current operation. I have found that a new HBO100W/2 lamp warmed-up and operated at 5 amps with a good DC supply has an arc voltage of about 14.8V, giving only about 75W. At 200 hours of life, this lamp had 28V across the arc (-at- 5 amps) and this is 140W! No doubt this relationship would continue until a) the lamp gets too hot and explodes, b) the power supply operating voltage becomes insufficient to maintain 5A current (giving a decrease in power and dimming of lamp output), or c) the power supply is unable to ignite and stabilize the lamp at the higher arc voltage. It is possible to operate the HBO100W/2 lamp at currents as low as 1 amp with the right equipment, although the light output is also way down. If you have a proper power supply and the light output is adequate, operating at less than the nominal rating should give a real extension of lamp life.
Another issue is the operating temperature of the lamp. Osram specifies that the lamp bases should be no higher than 230C degrees and that it SHOULD be operated at below 200C if possible. Lamp housings should be designed to provide sufficient heat dissipation at the nominal wattage by passive heat flow or air flow directed at the lamp bases: the quartz envelope must be hot to have stable Hg temperature and pressure but an excessive gradient along the lamp will produce excessive stress on the lamp. Clearly, with economy power supplies the power dissipation cannot be limited and at some point the lamp will get too hot and you risk catastrophic failure. More expensive electronic supplies allow for various sorts of automatic control or manual readout and adjustments to control or limit power levels.
In summary, there are very good reasons that modern equipment can give some lengthening of lamp life, but there are very real reasons not to push this too far: the lamps simply DO get consumed in the process of giving light. I have personally seen a 2" quartz collector shattered by a lamp explosion and that is quite an expense to replace.
Sincerely,
Dale A. Callaham Central Microscopy Facility University of Massachusetts Amherst, MA 01003 USA
} Date: Tue, 14 Nov 95 13:58:47 EST } From: "MARK DARUS, 8*346-2895, (216)266-2895" {darus-at-cle.dnet.ge.com} } X-To: mail11:; (-at-micro) } To: microscopy-at-sparc5.microscopy.com } Subject: sample charging
Mark, I assume that you are using an SEM and not a TEM. You did not say if you have tried using backscatter imaging to view your sample. It is not as effected by charging as secondary imaging and it will still give a good image. You can also try making a carbon paint trail from the stub to the top of the sample. This can make it easier for the electrons to get out of the sample.
Greg-at-umic.umic.sunysb.edu S.U.N.Y. at Stony Brook University Microscopy Imaging Center } } Does anyone have a "best" method for analyzing sections of } } quartz tubing, without it charging? I carbon coated the sample 3 times, } } I lowered the Kv, increased the strength of the condenser lens, increased } } the bias, increased the scan rate and tilted the stage 35-45 degrees. } } Still I get a lot of charging. When I can manage to get an area that is not } } charging, and am able to focus on it, my counts are very low, for EDX. The } } samples are sections of quartz tubing, that have either inclusions or } } devitrification that is tinted. Should I just continue adjusting the above } } parameters, or is there something else that I should try? } } } Thanks } } Mark Darus } } Darus-at-cle.dnet.ge.com }
There actually is a manual in microscopy that gets updated quarterly. It is put out by John Wiley and is called Procedures in Microscopy. It comes in a 3 ring binder so it can be easily updated. It is similar to the one in Molecular Biology if you are familiar with that. It has been out at least 2 years and IS updated regularly. Cheers, Judy M
Judy Murphy, PhD Dept. of Microscopy San Joaquin Delta College 5151 Pacific Ave Stockton, CA 95207
It seems to me that a manual of microscopy techniques would be extremely useful, better in several ways than a book. I've set out my thoughts on this in the following URLs (they contain identical information but access speed will be better from your closest site).
Europe - {URL:http://www.lars.bbsrc.ac.uk/micro/in-focus/9511.html} America - {URL:http://metro.turnpike.net/jefferie/in-focus/9511.html}
If the idea interests you, please take a look and let me have some comments. If there's enough response perhaps we could get a group of people discussing it more formally by e-mail to hammer out some consensus views.
------------------ RFC822 Header Follows ------------------ Received: by ms.sjdccd.cc.ca.us with SMTP;15 Nov 1995 10:21:59 -0800 Received: (from daemon-at-localhost) by Sparc5.Microscopy.Com (8.6.11/8.6.11) id SAA06388 for dist-Microscopy; Tue, 14 Nov 1995 18:24:27 -0600 Received: from stjohns.ohsu.edu (stjohns.ohsu.edu [137.53.1.80]) by Sparc5.Microscopy.Com (8.6.11/8.6.11) with SMTP id SAA06385 for {microscopy-at-Sparc5.Microscopy.Com} ; Tue, 14 Nov 1995 18:24:26 -0600 Received: by stjohns.ohsu.edu (Smail3.1.28.1 #18) id m0tFV8G-0007B5C; Tue, 14 Nov 95 15:51 PST Sender: kayton-at-ohsu.edu (Robert Kayton) Message-Version: 2 } To: microscopy-at-Sparc5.Microscopy.Com
We have an ISIS system and I would also like to hear whether anyone has successfully implemented Bioquant and, if so, what standards were used.
On Fri, 10 Nov 1995, Fiona Graham wrote:
} We are in the market for a new energy dispersive x-ray analysis system } to attach to a Hitachi S520 SEM. The systems we are considering } include: } } LINK ISIS } NORAN 3050 } EDAX DX4 } IRIDIUM II microanalysis system. } } If any current users of these systems have any comments about the } systems, we would most appreciate hearing from you. } } Thanks in advance } Dr Fiona Graham } EM Unit } University of Natal, Durban } South Africa } email: graham-at-ph.und.ac.za } tel: +27 31 2602174 fax: +27 31 2616550 }
MICROWAVE WORKSHOP 1996 WITH PRIMARY EMPHASIS ON 3-HOUR PROCESSING OF TISSUE FOR TRANSMISSION ELECTRON MICROSCOPY
SAN DIEGO STATE UNIVERSITY, SAN DIEGO CA JAN 25-27, 1996 Contact Steve Barlow Phone: (619) 594-4523 FAX: (619) 594-5676 email: sbarlow-at-sunstroke.sdsu.edu
SEE BELOW FOR ALTERNATIVE DATES/SITES
Routine processing of tissue for transmission electron microscopy in clinical and research applications is a time consuming process. This workshop focuses on the use of a laboratory microwave oven as a means to accelerate all aspects of tissue processing for TEM. The demonstrations, course handouts and "hands-on" experience will provide workshop participants with the tools necessary to rapidly assimilate a laboratory microwave into their protocols. Other areas covered in the workshop include microwave demonstrations of histological staining (periodic acid-methenamine silver stain of kidney basement membranes), immunostaining for TEM and digital image acquisition and printing. Each workshop is an intensive two and a half-day experience based on the following:
Giberson, R.T., Demaree, R.S., Jr. (1995) Microwave fixation: Understanding the variables to achieve rapid reproducible results. Micros. Res. & Tech (in press)
Giberson, R.T., Smith, R.L., Demaree, R.S. (1995) Three hour microwave tissue processing for transmission electron microscopy: From unfixed tissue to sections. Scanning 17, Suppl. V:26-27
Demaree, R.S., Jr., Giberson, R.T., Smith, R.L. (1995) Routine microwave polymerization of resins for transmission electron microscopy. Scanning 17, Suppl. V:25-26
"Hands-on" Microwave Workshop for TEM and Histology Calif. State University, Chico, June 15-17, 1995
COURSE OUTLINE, SAN DIEGO STATE UNIVERSITY
Jan 25, 1996 8:30-5:00 Lecture and microwave demonstration by R. T. Giberson
Demonstration of 3 hour microwave tissue processing
Jan 26, 1996 8:00-5:00 Participants will work in groups of 5 and process tissue using 3
hour protocol in laboratory microwaves
Jan 27, 1996 8:00-12:00 Microwave demonstrations of on-grid immunostaining, histological
staining, and digital imaging
Registration Fee for the SAN DIEGO WORKSHOP (workshop limited to 20 participants)
Workshop ONLY (includes the following) $500.00
1. Group dinner at the end of the first day 2. Lunch the first two days 3. Course handouts and all materials used in the workshop 4. Certificates of Completion mailed to each participant
Workshop AND 3 DAYS LODGING $700 at local hotel for 3 nights starting the night before the workshop
1. Group dinner at the end of the first day 2. Lunch the first two days 3. Course handouts and all materials used in the workshop 4. Certificates of Completion mailed to each participant
In the event a participant needs to cancel, written notification (fax is acceptable) is required. A $50.00 administrative fee will be charged for cancellations. 50% of the registration fee will be reimbursed for cancellations within 30 days of the workshop. Substitutions are acceptable.
ALTERNATIVE LOCATIONS AND DATES:
University of Maryland Dept. of Zoology, College Park, MD JAN 3-5, 1996 Contact Tim Maugel- Phone (301) 405-6898 FAX: (301) 314-9358 email: maugel-at-zool.umd.edu
Yale University School of Medicine New Haven CT JAN 10-12, 1996 Contact Paul Webster Phone: (203) 785-3219 FAX: (203) 785-7226 email: paul_webster-at-quickmail.yale.edu
NIH/Rocky Mountain Labs, Hamilton, MT JULY 1-3, 1996 Contact Dave Dorward Phone:(406) 363-9266 FAX: (406) 363-9204 email: dave_dorward-at-nih.gov
Madison Area Technical College, Madison, WI AUG 7-9, 1996 Contact Michael Kostrna Phone:(608) 246-6762 FAX: (608) 246-6880 email: msk5063-at-madison.tec.wi.us
--------------------------------------------------------------------------- ------------------------------------------------------------------------------ Dr. Steve Barlow EM Facility/Biology Dept. San Diego State University San Diego CA 92182-4614 phone: (619) 594-4523 fax: (619) 594-5676 email: sbarlow-at-sunstroke.sdsu.edu
} Does anyone know of a software which I can use to estimate the } connectivity of phases of interest ? I have some SEM binary images } showing pore distribution patterns, and would like to determine how well } the pores connected to each other, based on their connectivity values.
This was done extensively (and published) by Bob Ehrlich using erosion/dilation. The commercial verison is handled by Perception and Decision Systems, which can be reached by e-mail at Pads12-at-aol.com. If you do a literature search of the last 10 years (especially J. Sedimetary Petrology and AAPG Bull) you may turn up much more. Most generic image processing packages have erosion/dilation algorithms included.
A user in our central EM lab just crunched the electrode on our old analog pH meter. It was an Orion Ross electrode with a replacement cost of about $200.
Rather than just replace this expensive electrode on an old meter, I thought I would ask about what types of pH measurement are in use by other labs. A recent Fisher catalog has dozens of alternatives. Too many to pick just one without some advice.
If you have some ideas, let me know. I am looking for the best combination of cost, accuracy, ease of upkeep, and durability in a multiuser lab.
Jonathan Krupp Microscopy and Imaging Lab University of California Santa Cruz, CA 95064 (408) 459-2477 jmkrupp-at-cats.ucsc.edu
I'm confused regarding the discussion on Na vapor lamps. I believe the original discussion concerned the use of less expensive (home improvement store) lamps in darkroom safelights. Did the discussion somehow turn to the use of such lamps in fluorescent microscopes???? I think I must have missed a transitional message somewhere. Could some kind soul please update me on the status. Can cheaper Na vapor lamps be used in both situations. I'd be (pleasantly) surprized if they could. Many thanks.
############################################################################# John J. Bozzola, Director Center for Electron Microscopy 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 #############################################################################
In message Wed, 15 Nov 1995 11:01:10 -0800 (PST), sbarlow-at-sunstroke.sdsu.edu (Steve Barlow) writes:
} I'm looking for researchers using freeze substitution in their SEM and TEM } protocols. I know there are several nice insturments for this } protocol--I'ld like to contact researchers who are doing biological } freeze substitution without the benefit of these expensive devices and } who reside in the southwestern USA ----- The recent article on Freeze-substitution (Part I, Chapt.V) on Methods In Plant Cell Biology, Part A (Edts. Gailbraith et al.; Academic Press, 1995) might be helpful.
************************************************************************* M.V. Parthasarathy Professor of Plant Biology & Director, Cornell Integrated Microscopy Center Section of Plant Biology 228 Plant Science Building Cornell University, Ithaca, NY 14850 Telephone: (607) 255-1734 Fax: (607) 255-5407 E-mail: mvp2-at-cornell.edu ***********************************************************************
Thank you to all of those who sent me replies regarding "Monographs in Practical Electron Microscopy in Materials Science" by Edington. The 5 volume set has been combined into 1 volume and is available at a cost of $55 each. There is a 10% student discount available, and shipping costs (4.50 for 2 copies) are reasonable, at least in the U.S. There has been some doubt about the quality of the micrographs in parts, but I believe that they have to be better than the micrographs in the photocopies I have currently.
For more information/ To order contact:
Tech Books Inc 4012 Williamsburg Court Fairfax VA 22032 USA (800) 767-1518 (703) 352-0001 fax (703) 352 8862
The volume may also be available from
Polycrystal Book Service P.O. Box 3439 Dayton, OH 45401-3439 USA telephone and fax: (513) 223-9070
however they did not return my calls.
-Kirk ________________________________________________ Kirk A. Rogers krogers-at-materials.ecn.purdue.edu 317-494-8751 office http://materials.ecn.purdue.edu/~krogers 317-494-1204 FAX Purdue University, School of Materials Engineering, 1289 MSEE building, W. Lafayette, IN 47907-1289
} I'm looking for researchers using freeze substitution in their SEM and TEM } protocols. I know there are several nice insturments for this } protocol--I'ld like to contact researchers who are doing biological freeze } substitution without the benefit of these expensive devices and who reside } in the southwestern USA } } I'ld appreciate any help in this regard } } thanks in advance } } steve barlow } } } --------------------------------------------------------------------------- } ------------------------------------------------------------------------------ } Dr. Steve Barlow } EM Facility/Biology Dept. } San Diego State University } San Diego CA 92182-4614 } phone: (619) 594-4523 } fax: (619) 594-5676 } email: sbarlow-at-sunstroke.sdsu.edu } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } } }
The most critical part of freeze substitutuion is the freezing. After that there are a couple of low tech methods for substitution. : After freezing the sample cool to liq. nit. temp and place on the substitution medium that has been solidified by freezing in liq. nit. Then pack it in dry ice in a styrofoam box and wait for it to come to room temp.
I do the same only I place the samples in a -45 C freezer for two days then in a - 20C freezer for a day, instaed of on dry ice.
If there Are several freezers available you can place the frozen sample in - 85 C substitution medium and gradually raise the temp by moving the sample in the medium from freezer to freezer. -at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at- Greg Erdos Phone: 904-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax 904-846-0251 University of Florida E-mail: gwe-at-biotech.ufl.edu Gainesville, FL 32611 -at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at--at-
I don't know if your interest in freeze substitution is to actually try it out before buying a machine but if it is, here is a neat way to do it. Fix the biological material (either by freezing or by chemical means).
If you choose chemical fixation you must then cryoprotect the material by infiltrating with 2.0M sucrose and then freezing on metal pins, by dropping them in liquid nitrogen.
Once frozen, transfer the material to dry methanol (just open a new bottle) which is being held in a tube, in a styrofoam box filled with dry ice. Keep the material in the cold methanol overnight. Remove the methanol and replace with fresh methanol, leave for a few hours, and replace with a methanol-Lowicryl mix (1:1). If you want to keep the tubes on dry ice you can use Lowicryl HM 23, otherwise, transfer the tubes to a freezer. Keep increasing the amount of resin in the tubes until you are in 100% resin. Leave overnight, replace with fresh resin and polymerize with UV light. Great embedding (I have pictures) with low cost.
Variations include adding 1% osmium tetroxide or 1% uranyl acetate to the methanol. The contrast is subtle but significant and the osmium does not affect polymerization or immunolabeling.
Paul Webster Center for Cell Imaging Yale School of Medicine http://info.med.yale.edu/cellimg
But doesn't the Procedures in Microscopy manual cost } $500? Presumably an online manual maintained by people like us would be free.
} There actually is a manual in microscopy that gets updated quarterly. It is } put out by John Wiley and is called Procedures in Microscopy. It comes in a 3 } ring binder so it can be easily updated. It is similar to the one in } Molecular Biology if you are familiar with that. It has been out at least 2 } years and IS updated regularly. } Cheers, } Judy M } } Judy Murphy, PhD } Dept. of Microscopy } San Joaquin Delta College } 5151 Pacific Ave } Stockton, CA 95207 } } Phone: 209/474-5284 } FAX: 209/474-5649 } e-mail: murphy.ms.sjdccd.cc.ca.us } } _______________________________________________________________________________ } From: chris-at-stowey.demon.co.uk on Wed, Nov 15, 1995 10:22 AM } Subject: Microscopy Manual Online } To: microscopy-at-Sparc5.Microscopy.Com } } It seems to me that a manual of microscopy techniques would be } extremely useful, better in several ways than a book. I've set out } my thoughts on this in the following URLs (they contain identical } information but access speed will be better from your closest site). } } Europe - {URL:http://www.lars.bbsrc.ac.uk/micro/in-focus/9511.html} } America - {URL:http://metro.turnpike.net/jefferie/in-focus/9511.html} } } If the idea interests you, please take a look and let me have some } comments. If there's enough response perhaps we could get a group of } people discussing it more formally by e-mail to hammer out some } consensus views. } } Thanks, } } Chris } -- } ----------------------------------------------------------------------------- } | Chris Jefferies E-Mail (home) - chris-at-stowey.demon.co.uk | } | (work) - chris.jefferies-at-bbsrc.ac.uk | } | Microscopy page (UK) {URL: http://www.lars.bbsrc.ac.uk/micro/} | } | (USA) {URL: http://metro.turnpike.net/jefferie/} | } ----------------------------------------------------------------------------- } } ------------------ RFC822 Header Follows ------------------ } Received: by ms.sjdccd.cc.ca.us with SMTP;15 Nov 1995 10:21:59 -0800 } Received: (from daemon-at-localhost) by Sparc5.Microscopy.Com (8.6.11/8.6.11) id } SAA06388 for dist-Microscopy; Tue, 14 Nov 1995 18:24:27 -0600 } Received: from stjohns.ohsu.edu (stjohns.ohsu.edu [137.53.1.80]) by } Sparc5.Microscopy.Com (8.6.11/8.6.11) with SMTP id SAA06385 for } {microscopy-at-Sparc5.Microscopy.Com} ; Tue, 14 Nov 1995 18:24:26 -0600 } Received: by stjohns.ohsu.edu (Smail3.1.28.1 #18) } id m0tFV8G-0007B5C; Tue, 14 Nov 95 15:51 PST } Sender: kayton-at-ohsu.edu (Robert Kayton) } Message-Version: 2 } } To: microscopy-at-Sparc5.Microscopy.Com } From: !Microscopy-request-at-Sparc5.Microscopy.Com (Robert Kayton,MAC,CROET) } Date: Sat Oct 28 21:18:41 GMT 1995 } UA-Content-ID: {PMX-TERM-2.2-504424-stjohns-kayton-14662} } Received: from steele.ohsu.EDU by stjohns.ohsu.edu with smtp } Received: (Smail3.1.28.1 #18) id m0t9KHS-0007ASC; Sat, 28 Oct 95 16:03 PDT } Received: from Sparc5.Microscopy.Com by steele.ohsu.EDU (5.x/SMI-SVR4) } Received: id AA07353; Sat, 28 Oct 1995 16:12:49 -0700 } Received: (from daemon-at-localhost) by Sparc5.Microscopy.Com (8.6.11/8.6.11) id } QAA05400 for dist-Microscopy; Sat, 28 Oct 1995 16:19:32 -0500 } Received: from relay-4.mail.demon.net (relay-4.mail.demon.net [158.152.1.64]) } by Sparc5.Microscopy.Com (8.6.11/8.6.11) with SMTP id QAA05397 for } {microscopy-at-sparc5.microscopy.com} ; Sat, 28 Oct 1995 16:19:28 -0500 } Received: from post.demon.co.uk by relay-4.mail.demon.net id sg.aa02774; } Received: 28 Oct 95 21:17 GMT } Received: from relay-4.mail.demon.net by relay-3.mail.demon.net id aa11623; } Received: 28 Oct 95 21:17 GMT } Email-Version: 2 } Subject: Microscopy Manual Online } Reply-To: chris-at-stowey.demon.co.uk } Message-Id: {276-at-stowey.demon.co.uk} } To: microscopy-at-Sparc5.Microscopy.Com } X-Mailer: FIMail V0.9 } X-User: X-User: Alpha Test Version Of FI-Mail } Lines: 23 } Content-Type: text } Content-Length: 1115
Thomas E. Phillips, Ph.D. Associate Professor of Biological Sciences Director, Molecular Cytology Core Facility 3 Tucker Hall University of Missouri Columbia, MO 65211 (314)-882-4712 (voice) (314)-882-0123 (fax)
Microscopists, I am responsable for a small histology service lab. My new histologist has some experience with microwave stain techniques and use of microwaves to express antigens in archival-paraffin-embedded material. We are seriously considering the purchase of a microwave for our lab to speed things up, but we are wondering about the merits of "off the shelf" microwaves (eg appliance stores) versus "technical" microwaves available from a number of reputable microscopy suppliers. There is a considerable price difference between the two types and I'd like to be educated on the relative merits of one versus the other (our needs are not likely to include EM processing, since there is a seperate lab in the Health Sciences Center for that).
Perhaps it would be best if you replied to me personally and I can then post a summary to the list at a later date.
A note to vendors: I appreciate your input, to save yourself paper & mailing costs, you might inquire if I already have some of your materials regarding microwaves before you send me any more.
Thanks, Doug ..................................................................... : Douglas W. Cromey, M.S. Dept. of Cell Biology & Anatomy : : Sr. Research Specialist University of Arizona : : (office: AHSC 4212A) P.O. Box 245044 : : (voice: 520-626-2824) Tucson, AZ 85724-5044 USA : : (FAX: 520-626-2097) (email: dcromey-at-ccit.arizona.edu) : :...................................................................:
I plan to buy a nebulizer to prepare dispersions of particles in the 1-10 micrometers range. I have seen various models in as many catalogs. Can anyone recommended a particular model? Is there one to avoid at all cost? Thanks in advance for your comments. MICHEL
Michel Deschuyteneer deschuyt-at-sbbio.be Scientist - Electron Microscopy Laboratory SmithKline Beecham Biologicals Rue de l'Institut, 89 B-1330 Rixensart Belgium Tel: +32-2-656 9290 Fax: +32-2-656 8113
I have recently been sent a Mohr processor to test to see how it works in our lab for film and paper as we must get rid of our darkroom. This note is about the negative processing as I haven't even got to the print processing yet. Specifically I am having a major problem with Ektapan and SO163 (see below for specifics)
NOTE: Most of our electron microscopes are analogue so we would like to go digital but cannot totally do that at this time.
The films we need to have processed are (all Kodak films) 4489 EM film (3 1/4 x 4 inch) SO163 EM film (twice the speed of 4489) 3 1/4 x 4 inch Ektapan ( a 4. x 5 sheet film) continuous tone film for making copy negs, copying pictures, light microscopy in B/W 4127 (4 x 5 sheet film) good continuous tone used for SEM (Our student's can't afford Polaroid) The last two films are panchromatic films.
This system allows one to choose the time one processes in each chemical i.e. Mohr developer and standard Kodak Rapid Fix. This unit has a 600 watt heater on it. It also has the extended time switch.
We are using a temperature of 88 degrees for the dev and fixer (when measuring it, the dev is actually 88.5 and the fix is 84.5)
We run several plastic cleaning sheets through the processor before processing the negatives.
To give an idea of the initial clearing time in Kodak rapid Fix by wet chemistry, 4489 clears in 20 s in fresh fix; SO163 clears in 37 sec in fresh fix.
Every session we have worked on the Mohr, we have also developed the negatives by standard Kodak recommended and tried and proven "wet" chemistry using the same Kodak fix that is in the processor. All of the "wet chemistry" negatives have come out fine, as expected.
RESULTS from Mohr 4489 Works fine at the 2 minute setting. dry to dry.
4127. Works fine at 3.5 min. setting.
Ektapan and SO163 are problem childs. I have tried the Ektapan and SO163 at the following min. settings on the Mohr machine 2, 2.5, 3, 4, 5, 5.5, 6, 6.5 2 + Extended time (14.5 actual min from dry to dry) 3 + Extended time 6.5 + Extended time (17.5 actual min from dry to dry) As a rule of thumb, each of the settings on the machine is divided by 4 to get how much time is actually spent in each place (developer, fix, wash, dry) Thus the 17.5 actual time is divided by 4, which means 4.4 sec in each place. The 2 min setting means 30 sec in each place.
Because the first unit I got, the heater was not functionikng, I did my first tests without a heater. All had to be put in a dryer for about 2 min. to completely dry which they were. With and without a heater, 4489 and 4127 worked in the same times as indicated above.
Without a heater Ektapan, 2+Extended Time On (14.5 min. total) appeared to clear the photos, but wet. SO163, 5+Extended Time On appeared to clear the photos, but wet.
Heater arrives 4489 and SO163 Did the same tests with all the times, and at all of the times from the shortest 2.0 (Extended Time OFF) to 6.5+Extended Time ON, it not only didn't clear it but it precipiated black stuff (which I think is silver) all over the negatives in a potchy distribution. This could not be cleared anymore in fix applied outside the Mohr machine or washing outside. It was fried onto the negs. (Again 4489 worked fine)
I did have fresh fixer and developer. When I ran 4489 in between the Ektapan and SO163 tests, it still turned out at 2 min. (Ext. Time OFF).
I then removed the wash rack and ran negatives through the Mohr dev and Rapid fix (in the Mohr machine) and let them drop in the water bath. I did two things A. One neg I washed for 30 min. outside the Mohr to be sure it was washed, then ran it through the wash and dryer roller. The same thing happened. Stuff precipitated all over the neg. and fried on. B. One neg I removed after it went through the fix rollers and fixed it outside, followed by a fix neutralizer and washed it for 20 min outside. I then ran it through the Mohr wash rollers and dry. It also had stuff on it, fried to the neg as all of the others, but not quite as bad as those completely processed in the Mohr. I was very surprised by this results as I expected that if the unexposed Ag was completely washed out, it shouldn't happen.
Mohr doesn't have any suggestions and Ted Pella Inc. hasn't heard of this happening and has no suggestion. To add to all of this, my darkroom will very shorty be converted into a room to accept another instrument and I have no way of developing negatives unless I get a processor to do so. To say the least, my level of frustration is quite high. I have now spent about 16 hrs. testing this machine in a very methodical way, and still can't get Ektapan or SO163 to work. If there are any suggestions or similar experiences, I would really appreciate them. I really am out of things to try or do. Thanks Judy Murphy
Judy Murphy, PhD Dept. of Microscopy San Joaquin Delta College 5151 Pacific Ave Stockton, CA 95207
Mark, Sometimes (most of the time??) carbon is not an effective coating for neutralizing charge build-up on samples, biological in particular. Plus you get a noisy image so image signal quality is not great for photo recording.
Sometimes I've had to coat with metals in order to get a stable sample for doing x-ray microanalysis. The trick is to pick a metal that won't overlap with any elemental peaks collected from the sample, and to use as little as possible to minimize absorption of 0-2.0 KeV x-rays coming from your sample. For example, I'll use nickel or chromium or aluminum - just a little bit! - and often it will solve the charging problem so I have the same incident kV of primary electrons, always an important beam condition to stabilize for x-ray microanalysis.
Good luck,
--
Gib Ahlstrand, MMS Newsletter Editor Electron Optical Facility, University of Minnesota, Dept. Plant Pathology 495 Borlaug Hall, St. Paul, MN 55108 (612)625-8249 612-625-9728 FAX, giba-at-puccini.crl.umn.edu
"MICROSCOPY & MICROANALYSIS 96" in Minneapolis, Minnesota, Aug.11-15, 1996
Hi all, does anyone have experience with CY-5 on a "standard" HBO 50 lamp based Fl microscope. I've heard that it can be difficult to see (as opposed to detect with pmt's), the local sales droids have never sold the appropriate filter block so wont commit themselves. Dont particularly wish to fork out for the filter block if I wont be able see my results...
Anyone with advise/experience.
ta muchly --
Daryl Webb (dwebb-at-waite.adelaide.edu.au) Dept. of Plant Science, Waite Institute University of Adelaide, Glen Osmond S.A. 5064 Australia. Voice:61_8 303 7426 Fax:61_8 303 7102
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----- Unsent message follows ----- Received: from macmail7.lbl.gov by lbl.gov (4.1/1.39) id AA08019; Thu, 16 Nov 95 12:16:56 PST Message-Id: {n1395598242.52342-at-macmail7.lbl.gov}
Reply to: RE} Laser printing
to: Jean Leclef I am very interested in your message. Do you mean that this board can convert our laserjet 4+ to print grayscale instead of halftone (dithering)? Does the board go in the printer or in the host computer? Is a host a Mac or a PC or a Sun or a HP? Mike O'Keefe --------------------------------------
for those who are intested in laser printing
the following company in New York has a very good alternative to print images on Laserjet 3/4/4+ printers - it is a German interface board with 150, 300 and even 600 dpi printing capability, with a palette of 256 from 1024 gray levels. it works really fine for printing hires SEM images and prints a 2 MByte image in seconds!
the experience shows that it is a real alternative to photographic images when you have to find another way to print good pictures from a SEM/STEM.
contact : Electro Image, Inc. 9, Round Hill Road LAKE SUCCESS, NEW YORK 11020 tel 516 773 4305 fax 516 773 2955
I understand that charging up of samples is always a very problematic item in SEM. However under certain conditions of your sample and also depending on required resolution and magnification, LV-SEM (low vacuum SEM) can give a solution. Even under low vacuum condition excellent X-ray microanalysis can be done even on light elements.
If you want more detailed please contact us, or your nearest JEOL office.
Best regards.
Bob Hertsens JEOL Europe BV Ikaroslaan 7A B-1930 Zaventem Belgium
Two laboratory technician positions are available in a University analytical laboratory containing electron microscopes, optical scopes, a Rutherford backscattering spectrometer, X-ray diffractometers, and scanned-probe microscopes.
One position is at the Junior Lab Technician level and the other is a Senior Lab Technician. The duties will include assisting researchers with sample preparation, maintaining equipment, and assisting users from the University and from industry. The positions also include routine start-up and shutdown of instruments, maintaining and distributing supplies, and miscellaneous daily tasks.
Please contact me directly for further information.
_______________________________________________________________________ Beth Trend trend-at-cems.umn.edu Coordinator, Characterization Facility University of Minnesota Center for Interfacial Engineering 100 Union St SE Room 187 phone 612-624-1365 fax 612-626-7530 Minneapolis, MN 55455
Lots of people run them much longer than the specified time. There are potentially two big problems here:
1. After the life of the bulb is reached, the bulb starts a progressive drop in intensity. The first wavelength affected is the UV and it creeps up from there. Most people using FITC/Rhodamine, don't notice as much until the bulb is affected in their wavelength (400+ hours). If you are doing quantitative work, you will definitely want to change bulbs at the specified times.
2. Stability. The longer the bulb is run, the more the electrode and anode are worn. As they wear, the arc becomes unstable and dances on the ends. This causes flickering and some noise that can interfere with patch clamping. If a bulb explodes in your lamphouse, you will find that you now have to replace the collector lens, relay optics and sometimes the heat shields and mirrors. The biggest cause of exploding bulbs is on/off witching too frequently. A mercury bulb must be allowed to completely cool before be turned back on. A xenon bulb can be refired while still hot. To prolong the life of a bulb, it technically should be left on for long periods of time.
It should be noted that Osram has just come out with a new HBO 103 bulb which is the new replacement for the HBO 100W and is now rated to 300 hours for about $15-20 more than the old bulb.
Ted Pella, Inc. (PELCO #14601, Tel.#800-237-3526) has always supplied a hand-made glass one which I have used in the past for negative stained preps with excellent results. It isn't cheap but I think it's the best I've seen. BTW, I am in no way associated with this company and have not been solicited by Ted Pella, Inc.in any way.
Dan
On Thu, 16 Nov 1995, Michel DESCHUYTENEER wrote:
} Date: Thu, 16 Nov 1995 14:02:48 +0000 } From: Michel DESCHUYTENEER {deschuyt-at-ccmailg1.sbbio.be} } To: Microscopy-at-Sparc5.Microscopy.Com } Subject: SEM: nebulizer for particles dispersion } } Fellow microscopists: } } I plan to buy a nebulizer to prepare dispersions of particles in the } 1-10 micrometers range. } I have seen various models in as many catalogs. } Can anyone recommended a particular model? Is there one to avoid at } all cost? } Thanks in advance for your comments. } MICHEL } } Michel Deschuyteneer deschuyt-at-sbbio.be } Scientist - Electron Microscopy Laboratory } SmithKline Beecham Biologicals } Rue de l'Institut, 89 B-1330 Rixensart Belgium } Tel: +32-2-656 9290 Fax: +32-2-656 8113 }
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%% % % % Daniel Possin Work: 206/ 543-7489 % % Dept. of Ophthalmology RJ-10 FAX: 206/ 543-4414 % % University of Washington Home: 206/ 778-1714 % % Seattle WA 98195 USA Email: oemlab-at-u.washington.edu % % % % "The chinese expression 'cheung meng ba sui, gong hey fat choy' is % % equilvalent to Vulcan expression 'live long and prosper'. It's a % % small universe and getting smaller everyday". % % % %%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
I have encountered an annoying roadblock to getting our Kevex PDP-11/73 minicomputer on the network. For those familiar with this task: I have been
following John Mansfield's documented roadmap for getting these older EDS systems on ethernet. I've had no problem installing DELQA ethernet card in the PDP-11/73, but getting the software on our system has been an unexpected hassle.
The only input into our Kevex box is by way of an Iomega Bernoulli dual drive, which handles 44MB cartridges. Process Software, the company which makes TCPware for these computers, can't supply their software on this media, but instead offers 5-1/4 inch RX50 format or 8 inch RX01 format. Kevex Corporation has graciously agreed to copy the software onto our 44MB cartridges, but they can only accept 8 inch RX02 double density format, which Process Software is unable to provide. So, I am sitting here with the software we need, but apparently with no way of getting it onto the Kevex...
I have been reading up in my 'DEC microcomputer interfaces handbook' (very dry and nearly unintelligible text) and it seems that RX01 and RX02 drives differ only in the available bit densities. The RX01 drive has a bit density of 3200 bits/inch at the inner track, whereas RX02 runs at either 6400 bpi (double density) or 3200 bpi (single density, essentially equivalent to RX01). The handbook makes it very clear (ha ha) that the RX02 drive can be configured to write in single or double density mode, so I am assuming that Kevex's drive writes double density. However, it also says that when the drive reads, it determines the density of the media, which I interpret as the drive is able to read either density without a configuration change. In a private communication from Nestor, he also confirms that DEC RX02 drives and the DEC driver software can read RX01 floppies, and is curious whether or not Kevex understands their system.
In light of this information, I have contacted Kevex again and discussed this. They are willing to give the RX01 floppy a try, but they 'aren't sure if it will work' because they've 'never done this before.' Is there any real reason to believe that it wouldn't work? Isn't Process Software supplying their product on RX01 media so as to not handicap those who don't have an RX02 drive, since RX02 drives can handle both densities? Is anyone out there familiar enough with these types of data transfer such that they can counsel Kevex or instruct them on the correct RT-11 commands to perform the task? I find it hard to believe that it is more complicated than 'copy dy1:*.* dl1:*.*'....
Does anyone know of another way (besides Kermit) to get this software loaded onto the 44MB cartridges, perhaps through a 3rd party or something? The people at Process Software have been very helpful and are actively pursuing a solution, and they suggested trying the general microscopy community as well.
Thanks to all in advance for your advice,
Mike Mallamaci
_________________________________________________________ Michael P. Mallamaci mallamaci-at-goodyear.com Goodyear Tire & Rubber Company Tel: (216) 796-7436 D/415A Analytical Sciences Fax: (216) 796-3304 142 Goodyear Blvd Akron, OH 44305 USA
We also substitute in fixative/methanol or acetone on dry ice, but where possible leave the sample on dry ice for 2 weeks or so, with the intention that the substitution should happen at dry ice temp, not during warm-up. ---------------------------------------------------------------------- Sally Stowe |Email: stowe-at-rsbs-central.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, or 249 3808 |AUSTRALIA 0200
Message-Id: {9511172119.AA19829-at-MIT.MIT.EDU} To: microscopy-at-Sparc5.Microscopy.Com
This is really a very good point, anything put together by users as I propose would be free, easy to update, free of restrictive copyright, and you'd always see the latest version when you checked it for a method.
} But doesn't the Procedures in Microscopy manual cost } $500? Presumably an } online manual maintained by people like us would be free. } } } There actually is a manual in microscopy that gets updated quarterly. It is } } put out by John Wiley and is called Procedures in Microscopy. It comes in a } } ring binder so it can be easily updated. It is similar to the one in
I=B4m looking for good books (or online-manuals) on microscopical photography. Topics: theory, gradation, materials, effects of light on halogenides, applications in LM, TEM, SEM, comparisons to digital systems...
Any good hints available? Thanks in advance, -Dietmar-
+++ Dietmar Reiter {dietmar.reiter-at-uibk.ac.at} ++++++++++++++++ +++ Dept. of Zoology and Limnology, University of Innsbruck ++++ +++ Technikerstrasse 25, A - 6020 Innsbruck, Austria +++++++++ +++ ph: (+43)-512-507-6170 (-6161), fax: ..-507-2930 (-2957) +++
... If I would have had more time, I would have written a shorter e-mail ...
For those interested in the continuing and hopefully the ending saga of the Mohr processor problems reported by Judy Murphy, here is the problem, and results from our rather extensive by now, tests on films.
I am putting the original message on the bottom so if you haven't seen that but are interested, you might want to read it. (then again, you might not!)
After I had tried all the tests and removed the different sets of rollers and tried a mixture of hand chemistry and processor (as described in the original message, below), the rollers by then were dirty from whatever was being "fried" onto the surface of the Ektapan (4162) and SO 163. Also by that time I had gone through at least 150 sheets of film which is the limit for the solutions so had to change solutions as well. I removed the set of washer rollers, which is rather large) and put them in cleaning solution overnight as there was stuff baked all over them. The other sets of rollers I soaked in water. The next morning, with brush in hand, ready to scrub the rollers as this stuff was truly "baked" on, I turned over the washer rollers. and found that there were two pieces of 8 x 10 paper interwoven in the inside set of rollers. Thus certain parts of the rollers were not working. Some of them HAD to be working however, as 4489 and 4127 was being processed OK and all of the film was being transported properly out of the processor. It turns out that there is no easy way for the user to remove these sheets as there are abundant gears etc. which would require some expertise to get all the parts back together again. That is a bit of a worry to me, in that, if we start to depend on the processor and get rid of our chemicals, if the rollers jam, we are DOWN. We have 80 students printing and producing negatives, if we are down and can't process or print, our instruction programs comes to a grinding halt - all due to a set of rollers. Something for everyone to think about. Anyway on with the saga. I started in my testing all over again with the new set of rollers that were sent. Mohr shipped them overnight delivery, which was helpful. All of the films worked. It was just a matter of finding the correct times and re-doing everything I had done 2 or 3 times before. I continued to time the unit from leading edge in to leading edge out. Previously the setting of 2.0 (Extended switch ON) was 14.5 min. With the new rollars the setting of 2.0 + ext time ON (which should have been longer) was now 8 min. 3.0 + Ext Time ON was 10 min. It was obvious that the films were spending more time in the washer/dryer rollers then they should have. They might have even stopped at some point. That was unclear. I did not have time to test if all the extended times were now reproducible but the one on 3.0 minutes was. I did test that with the extended time OFF, the 2.0, 2.5, 3.0, 3.5, 4.0, 5.0, 5.5, 6.0 and 6.5 were as they stated.
Times that worked in the trial unit 4489 setting 2.0, Ext Time, OFF 4127 setting 3.5, Ext Time, OFF Ektapan, 4162, setting 6.5, Ext Time, OFF (see note below) SO163 setting 3.0, Ext Time, ON
The 600 watt is definitely necessaary for both SO163 and Ektapan. NOTE: At the above setting for Ektapan, i.e. 6.5 (Ext time, OFF), the film when it comes out of the processor, is just slightly, very slightly tacky and requires about 20 sec of RT to dry. Because we have a multi-user, multi-film facility, I am going to put the processor to the test with the Ektapan at 6.5 (with Ext Time OFF), so students don't have to constantly turn the ext time on and off. OTHERWISE, it is likely the extended time will be left on when it should be off and vice versa. When we process SO163, of course, we will have to use the extended time ON.
It is still unclear to me why the films previously were not being fixed properly even at the longest times, and now they appear to be. Perhaps there still IS unexposed AgBr in the films and it is not that obvious because it is not being baked which certainly made it visible. Definitely the SO163 has a small amount of stuff left as there is a slight haze on the film. It would be a good idea to get a Hypo Kit which tests to see if there is any unremoved material left in the films. Someone mentioned to me about this kit but I have never seen it. If anyone knows a source, please let me know. If there is material left in the films then of course the archival quality of the films is in question.
My suggestion for anyone buying a unit based on what I have had to do. Ask that the company process Ektapan 4162 on the particular unit they are about to send you. Ask also that the processed Ektapan be sent with the unit so you can see them. If 4489 is processed and works there is no guarantee that the washer rollers are actually working as was demonstrated in the unit I have. I spent copious hours in the darkroom and went through a great amount of solutions, film and most importantly time when the apparent culprit was the washer rollers. Again, that doesn't explain the lack of fix problem previously explained in the original message. That is still not clear to me. I hope they will now leave the unit long enough that we can actually use it to see if it will work in our multi-user facility that does both lots of film and prints. At the moment I only know what the times are and have worked out the problems with the unit. The logistics of doing both film and prints with so many students on the processor is yet to be answered.
Hope some of this has been helpful to someone. Cheers, Judy Murphy
Original Message I have recently been sent a Mohr processor to test to see how it works in our lab for film and paper as we must get rid of our darkroom. This note is about the negative processing as I haven't even got to the print processing yet. Specifically I am having a major problem with Ektapan and SO163 (see below for specifics)
NOTE: Most of our electron microscopes are analogue so we would like to go digital but cannot totally do that at this time.
The films we need to have processed are (all Kodak films) 4489 EM film (3 1/4 x 4 inch) SO163 EM film (twice the speed of 4489) 3 1/4 x 4 inch Ektapan ( a 4. x 5 sheet film) continuous tone film for making copy negs, copying pictures, light microscopy in B/W 4127 (4 x 5 sheet film) good continuous tone used for SEM (Our student's can't afford Polaroid) The last two films are panchromatic films.
This system allows one to choose the time one processes in each chemical i.e. Mohr developer and standard Kodak Rapid Fix. This unit has a 600 watt heater on it. It also has the extended time switch.
We are using a temperature of 88 degrees for the dev and fixer (when measuring it, the dev is actually 88.5 and the fix is 84.5)
We run several plastic cleaning sheets through the processor before processing the negatives.
To give an idea of the initial clearing time in Kodak rapid Fix by wet chemistry, 4489 clears in 20 s in fresh fix; SO163 clears in 37 sec in fresh fix.
Every session we have worked on the Mohr, we have also developed the negatives by standard Kodak recommended and tried and proven "wet" chemistry using the same Kodak fix that is in the processor. All of the "wet chemistry" negatives have come out fine, as expected.
RESULTS from Mohr 4489 Works fine at the 2 minute setting. dry to dry.
4127. Works fine at 3.5 min. setting.
Ektapan and SO163 are problem childs. I have tried the Ektapan and SO163 at the following min. settings on the Mohr machine 2, 2.5, 3, 4, 5, 5.5, 6, 6.5 2 + Extended time (14.5 actual min from dry to dry) 3 + Extended time 6.5 + Extended time (17.5 actual min from dry to dry) As a rule of thumb, each of the settings on the machine is divided by 4 to get how much time is actually spent in each place (developer, fix, wash, dry) Thus the 17.5 actual time is divided by 4, which means 4.4 sec in each place. The 2 min setting means 30 sec in each place.
Because the first unit I got, the heater was not functionikng, I did my first tests without a heater. All had to be put in a dryer for about 2 min. to completely dry which they were. With and without a heater, 4489 and 4127 worked in the same times as indicated above.
Without a heater Ektapan, 2+Extended Time On (14.5 min. total) appeared to clear the photos, but wet. SO163, 5+Extended Time On appeared to clear the photos, but wet.
Heater arrives 4489 and SO163 Did the same tests with all the times, and at all of the times from the shortest 2.0 (Extended Time OFF) to 6.5+Extended Time ON, it not only didn't clear it but it precipiated black stuff (which I think is silver) all over the negatives in a potchy distribution. This could not be cleared anymore in fix applied outside the Mohr machine or washing outside. It was fried onto the negs. (Again 4489 worked fine)
I did have fresh fixer and developer. When I ran 4489 in between the Ektapan and SO163 tests, it still turned out at 2 min. (Ext. Time OFF).
I then removed the wash rack and ran negatives through the Mohr dev and Rapid fix (in the Mohr machine) and let them drop in the water bath. I did two things A. One neg I washed for 30 min. outside the Mohr to be sure it was washed, then ran it through the wash and dryer roller. The same thing happened. Stuff precipitated all over the neg. and fried on. B. One neg I removed after it went through the fix rollers and fixed it outside, followed by a fix neutralizer and washed it for 20 min outside. I then ran it through the Mohr wash rollers and dry. It also had stuff on it, fried to the neg as all of the others, but not quite as bad as those completely processed in the Mohr. I was very surprised by this results as I expected that if the unexposed Ag was completely washed out, it shouldn't happen.
Mohr doesn't have any suggestions and Ted Pella Inc. hasn't heard of this happening and has no suggestion. To add to all of this, my darkroom will very shorty be converted into a room to accept another instrument and I have no way of developing negatives unless I get a processor to do so. To say the least, my level of frustration is quite high. I have now spent about 16 hrs. testing this machine in a very methodical way, and still can't get Ektapan or SO163 to work. If there are any suggestions or similar experiences, I would really appreciate them. I really am out of things to try or do. Thanks Judy Murphy
Judy Murphy, PhD Dept. of Microscopy San Joaquin Delta College 5151 Pacific Ave Stockton, CA 95207
Hey Daryl -- I have had good results visualizing Cy5 with a 100W HBO lamp on a standard epi-fluor. 'scope using Zeiss' "00" filterset (excitation: BP 530-585, beamsplitter: FT 600, emission: LP615) which is a filterset designed for visualizing Texas Red. However, you can't see cy5 fluorescence by eye. If you have a CCD camera you can use it to capture the Cy5 image as long as the CCD has decent spectral response above ~650nm. Many CCD cameras have an IR cut off filter so that they will match human vision. For the camera I use (Dage 72) it was a trivial matter to remove the IR cut off filter. Good Luck,
Greg Martin Dept. Cell Biology and Anatomy Johns Hopkins School of Medicine
On Fri, 17 Nov 1995, Daryl Webb wrote:
} Hi all, does anyone have experience with CY-5 on a "standard" HBO 50 lamp } based Fl microscope. I've heard that it can be difficult to see (as opposed } to detect with pmt's), the local sales droids have never sold the appropriate } filter block so wont commit themselves. Dont particularly wish to fork out } for the filter block if I wont be able see my results... } } Anyone with advise/experience. } } ta muchly } -- } } Daryl Webb (dwebb-at-waite.adelaide.edu.au) } Dept. of Plant Science, Waite Institute } University of Adelaide, Glen Osmond S.A. 5064 } Australia. Voice:61_8 303 7426 Fax:61_8 303 7102 } } } }
In regards to a hot debate we are currently having here, I am curious what other universities do about teaching TEM courses. My specific questions are:
1. How often do you teach a TEM course? 2. How many grad/undergrads take it? 3. Do you charge for consumables or beam time? 4. Does it make use of a departmental or campus wide facility? If so, does it interfere with researchers using the facility?
If you e-mail me directly I will be happy to post a summary of the responses to the list once they are all in.
Thanks for your input.
Thomas E. Phillips, Ph.D. Associate Professor of Biological Sciences Director, Molecular Cytology Core Facility 3 Tucker Hall University of Missouri Columbia, MO 65211 (314)-882-4712 (voice) (314)-882-0123 (fax)
I saw your response to Mike D.'s query. Could you post the phone number/address for Deltavision please? Someone here has the Scanalytics package, but it's too slow for our needs. Is the Deltavision as slow as Scanalytics to deconvolve?
A company called AutoQuant Imaging in Albany, NY, is beta testing a deconvolution package (3-D) called AutoDeblur-TLB which is based on Tim Holmes' mathematics. (phone 518.276.2138; aqisales-at-aqi.com)
-- Nancy L Desmond, Ph.D. nld-at-virginia.edu Department of Neurosurgery 804.924.5607 (voice) 804.982.3829 (fax) University of Virginia Health Sciences Center, Box 420 Charlottesville, VA 22908
Some advice on membrane filter papers would be greatly appreciated from your group. An honours project is being completed at Monash University, Clayton Campus using SEM and TEM procedures. Could you please suggest a membrane filter which has the following characteristics. -not dissolved in ethanol (fixing procedure) -is compatible with spurrs resin (or other eg. epoxy resin) -if the membrane is not transparent, it must be able to be dissolved eg acetone (will have seaweed zygotes grown onto the surface of the membrane, so nothing to toxic -pore size of around 10 microns(not crucial) -membrane filter diameter between roughly between 4cm and 10cm
I have approached Millipore who have not been able to suggest anything diffinite (possibly polycarbonate membranes which are transparent and so do not need to be dissolved). If you are aware of a compatible membrane type or a further contact person who might be able to help, it would be greatly appreciated.
Thank-you Mugette Marelic c/o Monash University Clayton Department of Ecology and Evolutionary Biology
We setup a new Internet WWW site (URL) for technical data and information about Crystals, Optics and Lasers. Any comments? Anyone know discussion grups about laser display and/or laser spectroscopy? Thanks.
Hi Christian, there is an excellent and versatile freeware Image analysis program (NIH-Image) available from zippy.nimh.nih.gov/pub/nih-image. It runs natively on macs and powermacs, or apparently under emulation using Executor on an IBM. There is a list similar to this one that covers uses of the program and general image analysis run by the university of minnesota, details of subscription are included in the about nih-image documentation available on zippy.
Ray
On Fri, 17 Nov 1995 CFILION-at-BRMVM1.VNET.IBM.COM wrote:
} Good morning, } } We want to performe precise measurements on samples } (i.e. hole diameter, distance from center to center etc..) } using our microscope. } } Is there a software that can performe this in an easy way. (i don't need to } relearn "C"). } } Any vendor can phone me now ! (on thos subject) } } } Christian Filion } Mat. eng. } 514-534-6602 } fax: 514-534-7310 } } Cfilion-at-BRMVM1.vnet.ibm.com }
I'm looking at serial sections of nerves on formvar (1% in chloroform) coated slot copper grids (pretty standard stuff). The grids are acetone and distilled water cleaned several times and coated as per standard (which has worked consistently well for 4 years). For the past few months I've been having considerable trouble with what appeared to be focus problems. It now appears that the film is "shifting". The film is carbon stabilised and clean; we've tried modifying formvar coating preparations, carbon coating and specimen storage, but the film movement is still present. It appears "tight" when on the grid, but appears to shift under the beam (as indicated by what appeared to be focussing problems). Anyone with a suggestion?
End Result:
We had considered most of the suggestions below already. We are now using 1 mm x 0.5 mm Cu slot grids pre-acid (glacial acetic for 30 seconds) treated, water rinsed (3X), acetone rinsed (habit), water rinsed (3X), blotted dry and touched on the dull side with a " coat quick G pen" (Daido Sangyo Co. Japan - what's in it I don't know, but...) and coated as per standard. This may be overkill, but it appears to be working well. Thanks to all who sent suggestions. Hope nobody minds me adding there name to their suggestions that follow...
Suggestions:
1. 1x2 mm slot grids are notoriously hard to completely stabilize even when carbon coated. If you are sure of your coating technique, I would begin viewing each new grid by spreading the electron beam just wider than the length of the grid slot and irradiate the whole thing for about 10-15 minutes prior to taking a closer look. This will strip the light elements out of the formvar film and "carborize" it making it more stabile. If this still is not enough I would consider making thicker films by increasing the concentration of your formvar solution. 1% Formvar in chloroform seems a little thin to me even with carbon coating. If you still have trouble, e-mail me again.
Daniel Possin
2. When I prepared grids as you described I used a formic acid step. By cleaning the grids for about 15 seconds in formic acid then rinsing well with distilled water the surface of the grid was better prepared to accept the formvar film. We also used the thicker Synaptec type of slot grid. After the formic acid the grids were cleaned as you described. Remember that the acid etches the grid so use it for only a short time.
Jack Megill
3. You might try "Grid Glue"
The recipe can be found under "Tips & Tricks" at http://www.biotech.ufl.edu/~emcl
Just click the Wizard
Greg Erdos
NB. There' a few suggestions on better binding of films / sections to grids on this WWW pg.
4. You might want to switch to another film made from Butvar. It is available from Electon Microscopy Sciences, Cat. #01185. A 0.25% or 0.3% solution in chloroform is used, and films are cast from slides onto water the same as is done with formvar.
Martin B. Garment
5. We have switched to using Butvar films on slot grids, and find that they are much more stable. It helps if you pre-treat the grids in a dilute solution of the butvar (0.1%), not to make a film, but just to get the metal coated (spread the grids on filter paper in a glass dish, just spray the grids with drops from a pipette, and let dry--use a fume hood!). Then do your regular filming procedure. Butvar is available from Electron Microscopy Sciences 321 Morris Road, Box 251, Fort Washington PA 19034, 800-523-5874, and they also have technical reprints available.
Evelyn Clausnitzer
6.
I would have several concerns. I presume that you are also using mesh grids periodically and finding no focus problems or specimen drift? Otherwise, you should consider the possibility of a microscope problem.
Regarding slot grids. Carbon stabilization is essential. Narrow slots (0.5mm x 2.0 mm) are more stable than wide slots (1.0 mm x 2.0 mm). If a batch of slots are acting unstable (too little carbon?; contaminated formvar solution giving films with holes or streaks?) one can try to reduce uneven beam damage and drift by beginning the grid exam at low power and far from crossover, previewing a wide portion of the grid while the formvar cooks out and hopefully becomes more stable. Then at high mag and closer to crossover, the beam is less likely to overheat the formvar locally and drifting will be reduced.
We cast formvar films from a 0.5% solution in ethylene dichloride. Depending on the weather, the same bottle may be good for several months before acting poorly: holes and streaks, if seen, are a warning that it is time to open a new stock bottle.We preview new batches of grids under the EM beam (2-3 grids per film) after carbon shadowing, but before section collection, to avoid those which are holey or streaked, as these are likely to be unstable.
Sorry that I don't have a means to exactly quantitate the carbon shadowing process. We recently published a chapter on serial section methods in Methods in Cell Biology, Vol. 48, Academic Press. H. Epstein and D. Shakes, Eds.
David H. Hall
7. We have found it really valuable to "review" internally some of the problems others seem to encounter. It makes us better here when it comes to doing our own in-house work and of course, it makes us better in giving expert customer technical service when one of our customers has difficulties.
Here is the synopsis of what my internal people have concluded:
1] Your specimen holder is not tight enough. Is that possible? I am sure you would have checked this, but this was #1 on the list of the people in our own in-house laboratory.
2] Sections are too thick, causing a heat buildup.
3] Has *anything* changed, for example, did you change bottles of Formvar? If yes, then that could be your problem. "Formvar" is a trade name (owned by Monsanto in the USA) and is not the "generic" term often times used. The "Formvar" family of resins consists of a broad number of different products, all the same polymer but with different molecular weights, etc. Only one or possibly two of these products have the right characteristics for this particular application. We know exact what is the correct product when we purchase it to fill our bottles. I know also that there are some EM suppliers who just don't understand this need to put into their product bottles the exactcorrect "Formvar" product.
4] The TEM grids you are using could be a problem also, since we know that from some source, the slotted grids are not as flat as they ought to be, causing the section to be position independent in terms of what is focus. To my way of viewing things, this variation should not be enough to cause a focussing problem, but others seem to believe other wise. Have you checked the grids in terms of their flatness?
You can take a look at the range of SPI products, including grids by looking us up on our site on the WWW, the URL being given below. Our products are now being distributed by Oxford Instruments in Australia and you can find the details for contacting them on our "Australia" agent page via our web site.
Let me know if I can be of any further assistance to you with regard to this particular problem (or some other one you might pose to us).
Chuck (Charles A Garber)
8. Your procedure has worked for 4 years (and you carbon coat the Formvar), so I would question the microscope. Dirty apertures maybe? We use Formvar coated slot grids (no carbon coating) all the time and deal with occasional drift by "ironing" the sections and Formvar with the beam. The person that comes up with something better than Formvar or Butvar will make many microscopists very happy! Anyway, I would check the scope.
Beth Richardson
9. I have two things you might think about. First, are the grids from the same batch that you were using in past years or are they new? There may be an adherence problem simply because of a different surface structure. Second, are you sure that the film is moving? What you describe could be an instability in the TEM's high voltage.
Russell E. Cook
10. Just a thought - are you sure it is the film and not another problem? We had something similar a long time ago which turned out to be the fixing of the grid in the specimen holder. There was some grease combined with a need to service the holder (Philips EM300) so the the spring was more 'grippy' onto the grid. If other workers, or non-filmed grids, do not have the problem then I guess it is the film (or HT problem?). Good luck
Keith Ryan
11. Is the movement only seen on slot grids or do you see on small mesh (200) also? The carbon stabilization should have done the trick and I am just wondering if the holder itself is unstable in some way. If focusing is the issue and not lateral movement, one VERY long shot could be the stability of the objective lens current. Sweeping the potentiometer knob repeatedly through the focus range can help clear dirt from the contacts and a shot of electrical contact cleaner behind the panel should be attempted before moving to more costly diagnostics. After all other low-cost options are exhausted, call for service.
Doug Davis
12. I have had good luck with specimens which shift under normal beam conditions by using very low beam currents (~10 pA). In order for this to give you a decent image, you need either to have a long exposure time or to use very sensitive film. LoDose or MRF32 will be sensitive enough, but they have large grain--there is still no free lunch. If you have a very sensitive video system, that will make focusing easier--we have an intensified CCD which gives a useful image at video rates, so focusing can be done in a few seconds. I suppose that using even a large mesh grid would not be satisfactory, but if it is, then that may also stabilize your specimen.
Bill Tivol
13. Maybe this will help. I had a similar struggle a while back, and it turned out to be the microscope. It was a new Joel, and the beam would charge on the grid and shift just when taking a picture, causing the photograph to look as if the tissue was drifting. We convinced Joel it was a microscope problem, and they fixed it.
John F. Smiley ______________________________________________
Shaun Sandow, Division of Neuroscience, John Curtin School of Medical Research, Australian National University, ACT 0200
In response to Daniel Possin's recommendation of the Ted Pella nebulizer, I have to agree (and I'm a competitor!). This device was designed by a microscopist, and is a product custom-built for this purpose. It is expensive because it is produced in small quantities by a large glassware company who normally make things in the tens of thousands. I suggest you contact Ted directly at teppel-at-snowcrest.net, as I believe he has publications available relevent to this product. Steven Slap, Energy Beam Sciences
If anyone can donate a used TEM (less than 10 years old) to be sent to Vietnam, please contact Dr. Judith Ladinsky, Director of International Health Affairs at UW-Madison. She can pay for shipping, and can be reached at: jlladins-at-facstaff.wisc.edu Martin B. Garment Ophthalmology and Visual Sciences 1300 University Ave. Rm. 6687 Madison WI 53706 Voice (608) 262-9596 Fax (608) 262-0479 Email - mgarment-at-facstaff.wisc.edu
Tony Garrat-Reed wrote: } Dear Netters, {snip} } The TIFF format is quite precisely defined. It originated } with Aldus Corp., and was supported actively by them for a while. } Later, as it became a *de facto* standard, Aldus's support became passive. } Since Adobe bought out Aldus, they have taken over the support. There } is lots about TIFF in the Adobe support forum on Compuserve, and I } expect that there are other places, too, but I don't know them. You } can download the complete TIFF 6.0 specification via anonymous ftp } from SGI.COM in directory graphics/tiff. The filename is tiff6.ps } (it is a postscript file, so should be downloaded as ASCII, and it } is big!). } I will have to check out the reference you just cited. But I will take some exception. TIFF has much flexibility/variability built into it. As a result, you might say that there is no such thing as a "standard" TIFF file. Programmers can choose to include certain fields and omit others. There is some redundancy in the spec so that the same information can be specified in different ways.
A robust TIFF reader should be able to handle any combination. However, we all deal with finite software written at some point in time. It may not be able to handle some combinations of fields. Some months back, I posted that I found PageMaker, Microsoft Word, Graphics Work Shop and some other apps dealt with the images differently. What worked on one app sometimes failed on another. Hopefully the current versions are more comprehensive.
To the best of my knowledge, the Mac-specific stuff gets added and deleted as necessary when shipped properly through an application like Fetch. I suspect difficulties were in the reader-apps on whatever end. ---------------------------------------------------- Warren E. Straszheim 270 MD Ames Lab/ISU, Ames IA, 50011 Phone: 515-294-8187 FAX: 515-294-3091 E-Mail: wes-at-ameslab.gov (or: wesaia-at-iastate.edu)
coal characterization and processing electron microscopy, x-ray analysis, image analysis computer applications
I would appreciate it if all would send their answers to the list server as I would like to see the original data as well as a summary (my cake and eat it too). thanks judy murphy _______________________________________________________________________________
In regards to a hot debate we are currently having here, I am curious what other universities do about teaching TEM courses. My specific questions are:
1. How often do you teach a TEM course? 2. How many grad/undergrads take it? 3. Do you charge for consumables or beam time? 4. Does it make use of a departmental or campus wide facility? If so, does it interfere with researchers using the facility?
If you e-mail me directly I will be happy to post a summary of the responses to the list once they are all in.
Thanks for your input.
Thomas E. Phillips, Ph.D. Associate Professor of Biological Sciences Director, Molecular Cytology Core Facility 3 Tucker Hall University of Missouri Columbia, MO 65211 (314)-882-4712 (voice) (314)-882-0123 (fax)
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} Can anyone tell me how safe sodium vapor lamps are for } autoradiography emulsions? Are special filters needed? } Thanks, } Dear Pat, If you are using an x-ray film emulsion, such as that on LoDose film, you must develop the film in total darkness. When I did autoradio- graphy, I used NS2T film and put it through an x-ray film processor, so total darkness was not too burdensome a condition. Yours, Bill Tivol
We use a Thomas Safelight w/ the filters they recommend for processing x-ray films. These filters are much less tranmissive than the normal Thomas black & white filters but seem to offer complete protection against any fogging. I have tested Kodak NTB-2 coated slides exposed for } 24 hours to light from this lamp compared with similar slides kept in a dark box for the same period and found no apparent difference in background grain numbers. You will have to contact Thomas for information on the x-ray type filter set (as I cannot locate currently locate mine). I hope that this helps.
Dan
On Thu, 16 Nov 1995, Pat Masarachia wrote:
} Date: Thu, 16 Nov 1995 16:28:03 EST } From: Pat Masarachia {pat_masarachia-at-Merck.Com} } To: Microscopy-at-Sparc5.Microscopy.Com } Subject: Sodium Vapor Lamp and Autoradiography } } Can anyone tell me how safe sodium vapor lamps are for } autoradiography emulsions? Are special filters needed? } Thanks, } } Pat Masarachia } Bone Biology } Merck Research Laboratories } West Point, PA 19486 } phone 215-652-7999 } e-mail pat_masarachia-at-merck.com } } }
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%% % % % Daniel Possin Work: 206/ 543-7489 % % Dept. of Ophthalmology RJ-10 FAX: 206/ 543-4414 % % University of Washington Home: 206/ 778-1714 % % Seattle WA 98195 USA Email: oemlab-at-u.washington.edu % % % % "The chinese expression 'cheung meng ba sui, gong hey fat choy' is % % equilvalent to Vulcan expression 'live long and prosper'. It's a % % small universe and getting smaller everyday". % % % %%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
Perhaps there is someone out there with some experience in obtaining sections of very tough plant materials that may have silica or other crystalline inclusions. I have been requested to help with a project of sectioning Phragmites stems and rhizomes and the information I received was that it has so many knife dulling inclusions that this has not yet been sucessfully accomplished. Of particular interest to us are the "pigment bodies" that are located in the cells of specimens that grow in saltwater or brackish areas.
I have recently seen a posting from Dr. Garber at SPI Supplies detailing the qualities of "materials sciences diamond knives" but wonder if anyone has other suggestions.
In advance I thank you for consideration,
Sincerely,
Dale A. Callaham Central Microscopy Facility University of Massachusetts Amherst, MA 01003 USA
contact: Marylou Montross, Astronomy Dept., SDSU telephone: (619)594-6182 Steve Barlow, Electron Microscope Facility, SDSU (619)594-4523
Open House, College of Sciences, San Diego State University
Inner Space, Outer Space
Visitors to San Diego State University will explore worlds invisible to the naked eye on Saturday, December 2, 1995, from 6-10 pm when the College of Sciences at SDSU hosts the second annual "Inner Space, Outer Space" Open House. Amateur scientists and the general public are welcome at no charge. Light refreshments and T-shirts will be on sale to raise money for undergraduate student scientist activities. Scientists and students of the EM Facility, the Geology Department and the Astronomy Department will display powerful microscopes, telescopes, and computers that offer views of the universe ranging from microscopic parts of a mammalian cell to satellite images of earth and worlds beyond our own planet. Visitors to the EM Facility in the lower level of the Physical Science building will see Inner Space-- intricate details of the surface and internal structures of plant and animal life magnified 1,000 to 50,000 times on the transmission and scanning electron microscopes. Students will explain the operation of the microscopes and demonstrate the same sample preparation techniques used to prepare tissue samples from biopsies at the area hospitals. Remote access from area classrooms to the electron microscope via Cox Cable TV or the Internet will also be demonstrated. On the rooftop of the adjacent Astronomy Department, several refracting and reflecting telescopes will be focused on Saturn, the moon, and other celestial bodies of Outer Space. The public is invited to an exciting planetarium show featuring San Diego's night sky. An astronomer will identify the heavenly bodies visible from your own back yard over the course of a year. Also on view will be a laserdisc show of images from the Voyager space probe as it passed by Saturn and Jupiter. Members of the Astronomy department will provide lively commentary. The Chemistry/Geology Computer Lab will exhibit satellite images of the earth as seen from Outer Space, as well as images from the Jupiter Galileo probe, which is scheduled to enter the atmosphere of Jupiter on December 7. Members of the Geology Department will discuss how satellite imaging helps us to understand our own world, as well as worlds far away. To reach the Inner Space, Outer Space displays, take route 8 to the College exit and turn south to San Diego State University. Free parking is available on the top level of the College Avenue Parking Structure. Take the pedestrian overpass across College Avenue and proceed right, to the Chemistry/Geology building. Follow the signs to the laboratories located in the adjacent Physics and Physical Science buildings. The Open House will take place rain or shine, as most of the exhibits are indoors.
For more information, call Marylou Montross at 594-6182.
--------------------------------------------------------------------------- ------------------------------------------------------------------------------ Dr. Steve Barlow EM Facility/Biology Dept. San Diego State University San Diego CA 92182-4614 phone: (619) 594-4523 fax: (619) 594-5676 email: sbarlow-at-sunstroke.sdsu.edu
Microscopists, Below is my original post regarding questions about microwave hardware. A summary of responses follows. Thanks to all who responded.
I checked with our safety dept. and they don't have a policy about microwaves (except that they be UL listed and unmodified). If a hazardous chemical is going to be "cooked", then safety expects that the microwave will be placed in a fume hood. The only problems that safety has had with microwaves is when people start the microwave and don't monitor it's progress (to view a WWW page that demonstrates what happens to a simple table grape when it's microwaved too long, go to: http://www.sci.tamucc.edu/~pmichaud/grape/).
Doug Cromey
-----original posting-----
Microscopists, I am responsable for a small histology service lab. My new histologist has some experience with microwave stain techniques and use of microwaves to express antigens in archival-paraffin-embedded material. We are seriously considering the purchase of a microwave for our lab to speed things up, but we are wondering about the merits of "off the shelf" microwaves (eg appliance stores) versus "technical" microwaves available from a number of reputable microscopy suppliers. There is a considerable price difference between the two types and I'd like to be educated on the relative merits of one versus the other (our needs are not likely to include EM processing, since there is a seperate lab in the Health Sciences Center for that).
Perhaps it would be best if you replied to me personally and I can then post a summary to the list at a later date.
A note to vendors: I appreciate your input, to save yourself paper & mailing costs, you might inquire if I already have some of your materials regarding microwaves before you send me any more.
Thanks, Doug ..................................................................... : Douglas W. Cromey, M.S. Dept. of Cell Biology & Anatomy : : Sr. Research Specialist University of Arizona : : (office: AHSC 4212A) P.O. Box 245044 : : (voice: 520-626-2824) Tucson, AZ 85724-5044 USA : : (FAX: 520-626-2097) (email: dcromey-at-ccit.arizona.edu) : :...................................................................:
-----response-----
Doug: We use a plain, kitchen-type mw for our processing (principally fixation for light microscopy and EM, and specifically, to enhance fixative penetration). We have had no problems, and have had really nice fixation (as long as the undergrad student doesn't forget the water ballast!) We have not checked for hot-spots (with a little rack of neon lamps), but use the center of the ovenfor our material). I'd call us happy with the "low tech" approach. But if I had a protocol that called for organic solvents, I wouldn't hestitateto use the lab-style. One good oven explosion could spoil your whole day... HTH Julian Smith III Biology Winthrop University
-----response-----
In response to Doug's letter, let me add that the Fire Safety Officer here at the Health Sciences Centre has disallowed our microwave oven for use in the fume hood because it's only a domestic grade, and he insists that it is a fire hazard because volatiles might get into the electronics, even though it is in a very high volume fume hood. I think perhaps that we are the only lab in the world that has this problem, and I'd like to hear from others (as many as possible) to prove to the fire safety people that the hazard is basically nil. Please let me know what the situation is in your labs.
Garry Burgess Charge Technologist - Electron Microscopy Health Sciences Centre Winnipeg, Canada
-----response-----
Hi Doug, We are am EM facility and we currently use an older GE. All you need to do is spend a day calibrating it and finding it,s various hot spots. The "Scientific" models are designed to reduce these hot spots but at a cost. A simple way to find them in both types is to place a sheet of styrofoam on the bottom, run the microvave, and map the places where it begins to melt. You can get moe of a 3 dimensional picture by elevating the styrofoam. I have also seen somewhere a calibration pad sold by some of the EM suppliers although I don't remember where. More modern micros have a more even heating pattern. I hope this helps.
} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} {} { Scott D. Whittaker 218 Carr Hall Research Assistant Gainesville, FL 32610 University Of Florida ph 904-392-1295 ICBR EM Core Lab fax 904-846-0251 sdw-at-biotech.ufl.edu http://www.biotech.ufl.edu/~emcl/
-----response-----
Doug- As you asked, I am a vendor responding to see if you have our material on our laboratory microwaves. For a general, and totally non-commercial, discussion of the merits of laboratory microwaves in general, see our home page: http://www.mwrn.com/ebs/ebs.htm
I also recommend the Microwave Cookbook for Microscopists. Steven Slap, Vice-President
-----response-----
Dear Doug, The key advantages of laboratory microwave ovens over domestic microwave ovens are: 1) safety features (such as a high volume exhaust) 2) enhanced programming features (such as microwave cycle time or duty cycle, heating/cooling cycles, more control over time and power function) 3) accessories (such as computer control and monitoring, water circulators, vaccuum processing)
There is also a common element between domestic and laboratory microwave ovens- they are both large cavity devices. The significance (regardless of advertising claims) is that microwaves are distributed nonuniformly in the cavity and in the specimen (e.g., slide sections, tissues).
There are several published methods to easily determine how uniform and how reproducible a particular microwave oven model is. In my experience, it is important to select a microwave oven based on its uniform distribution of microwave energy. The current literature in microwave-accelerated specimen preparation also places more emphasis on calibrating the microwave oven for the particular application than on the hardware. (Please contact me if you need this literature- 4 books, several articles)
Gary R. Login, D.M.D., D.M.Sc. Assistant Professor of Oral Pathology Beth Israel Hospital Department of Pathology 330 Brookline Avenue Boston, Massachusetts, 02215
You requested the following information about the Lehigh University Electron Microscopy Courses:
1. How often do you teach a TEM course? ==} Basic SEM and TEM undergraduate course taught in fall semester each year. Advanced TEM taught every other year in the spring semester, alternating with Advanced SEM and X-ray Microanalysis.
2. How many grad/undergrads take it? ==} 15-30 students take the undergrad SEM/TEM course, aand perhaps half of these are actually graduate students. About 10 graduate students take the Advanced TEM course and the Advanced SEM and X-ray Microanalysis course.
3. Do you charge for consumables or beam time? ==} Both are charged to the courses making them rather expensive to run since each course has approximately 10 different laboratories.
4. Does it make use of a departmental or campus-wide facility? If so, does it interfere with researchers using the facility? ==} All electron microscopes are in the campus-wide central facility located in the Dept. of Materials Science and Engineering. All departments including biology and geology use the instruments in this department. In addition to our six research electron optics instruments, we maintain three 1970s-vintage microscopes for use by our undergraduate and graduate students: a Philips EM301 TEM and two ETEC Autoscan SEMs.
CE Lyman
Charles E. Lyman Phone: (610) 758-4249 Prof. of Materials Science and Engineering Fax: (610) 758-4244 Lehigh University E-mail: cel1-at-lehigh.edu 5 East Packer Avenue Bethlehem, PA 18015-3195
I read with interest Tony Garratt-Reed's recent posting on TIFF files and certainly concur with his assertions. It seems that his note was in response to a question that I did not receive or accidentally deleted without reading. However, I would like to elaborate slightly on his comments relative to Mac's and TIFF files.
The TIFF format is precisely defined. However, my observation has been that applications to read TIFF files are not. This is especially true in the case of software produced by instrumentation vendors. Many vendors produce file formats that comply with the TIFF specification and they produce "limited" TIFF readers that can read these files. These same files, when read by a truly generic TIFF reader, will be read and interpreted. However, the inverse is not always true. It is likely that you will come across other TIFF files that also comply with the TIFF specification but can not be read by the same "limited" TIFF reader. I have encountered this problem when taking images from our SEM image analysis system to our LM image analysis system. The solution has been to import the original image into a program such as Adobe's Photoshop and then rewrite the image to a new file. Although this is not a universal solution, it usually produces a TIFF file that can be read by a wider range of TIFF readers. I do agree that the TIFF reader in NIH Image seems to be more robust than I have found in any other image analysis software. Relative to the file type and creator information added to a Mac file, I am not enough of a ResEdit user to know how this information is incorporated into the file. However, I would be very surprised if this information was included in the TIFF file header. My impression is that this information is included "externally". I say this because I have taken TIFF, Word, WordPerfect, and Excel files to and from Mac's and PC's with no problem. If the file type and creator information is organically imbedded in these files, I would not expect the file transfer to proceed as smoothly as it does.
Personally, I have had no problem bringing TIFF files into or out of a Mac. In fact, I use the Mac to get a usable TIFF file to bring to a number of IBM/PC applications. Between NIH Image and Photoshop, I have read every TIFF file that I have come across and in turn, wrote each to a PC format disk for export to an IBM/PC or clone. I have also used the Mac to convert TIFF files to JPEG or GIF images using Photoshop and Word for Word, respectively. I have exported these images to PC's and UNIX systems with no problem. An example of an image that was converted from TIFF To GIF can be seen on my home page (Yes, this is a cheap plug to get you to look at my home page that is still in a state of development, like everyone). As a side, I use the Access PC software for Mac to PC conversion. Although the PC Exchange software that comes with System 7.5 is supposed to do the same thing as Access PC, I haven't had universal success with it as compared to Access PC.
As for Fetch, I usually transfer files in the Raw Data format. I haven't had any problems with this approach. However, I only use Fetch going to and from UNIX and a Mac. I haven't sent or received any files that were to be used on a PC. I SneakerNet all of my PC images. Another nice aspect of Access PC is that it supports removable media hard drives such as Syquest.
Larry Sutter Michigan Technological University Dept. of Civil and Environmental Engineering 1400 Townsend Dr. Houghton, MI 49931
South Bay Technology will be hosting a VERY informal Tripod Polisher User's group meeting at the MRS Conference in Boston. The MRS meeting will be held from Nov 28 - Dec 1 at the Marriott at Copley Place.
The User's Group Meeting will be held in the South Bay Technology Booth on Thursday November 30. South Bay Technology's booth (booth #1) is located in the Atrium Lounge in the Marriott at Copley Place.
All interested parties are welcome! We'll look forward to seeing you there!
David Henriks TEL: 800-728-2233 (toll-free in USA) South Bay Technology, Inc. 714-492-2600 1120 Via Callejon FAX: 714-492-1499 San Clemente, CA 92673 USA e-mail: 73531.1344-at-compuserve.com sbt-at-msa.microscopy.com
Workshop Objective This course will cover all aspects of pre-thinning and focus on final thinning via Tripod Polishing. Due to the limited class size and the extensive hands-on opportuinities, this course is well suited to novices as well as advanced Tripodders. The course will include sections on:
How to do it and why should I? What's really going on and what am I really seeing? How to prepare small, specific area cross-sections. The problem of wildly differing materials (eg tungsten). Rapid preparation of TEM cross-sections. Preparation of a wide range of materials: semiconductors, ceramics, metals,...
Hands-on Opportunity This course will be unique in that it will provide a hands-on opportunity for every class participant. Tripod Polishers, Polishing Wheels, and pre-thinning equipment will be made available to all participants and actual samples will be prepared - by the students - as part of the course. This is a great opportunity to get your hands dirty and actually learn by doing. The instructors will walk you through each step of the process and then let you loose on the equipment. This course is designed to teach the Tripod Polishing technique. Silicon samples will be provided to the students and used as the basis for the course teaching.
Workshop Location and Dates South Bay Technology - San Clemente, CA Dates: Friday & Saturday - February 23-24, 1996
Previous Participants (partial list) INTEL, AMD, Motorola, LSI Logic, Conner Peripherals, Univ of Maryland, Univ of New Mexico, UNAM (Mexico), LG Electronics (Korea), Battelle, MEMC, MVA Inc., Univ of Michigan, U.S. Bureau of Mines.
Class Size Due to the intensive hands-on aspects of this course, class size will be strictly limited to 10 participants.
Registration Fee: $795 (includes lunches and Friday night Dinner) $695 if registration fee paid by January 12, 1996
Registration Deadline: January 30, 1996
For additional Information: Monica Pflaster South Bay Technology, Inc. 1120 Via Callejon San Clemente, CA 92673 TEL: 800-728-2233 FAX: 714-492-1499 e-mail: sbt-at-msa.microscopy.com
Registration Form
To register for the workshop, please fill out this form and send it, with registration fee, by January 26, 1995 to:
South Bay Technology, Inc. Workshop on Tripod Polishing 1120 Via Callejon San Clemente, CA 92673
Payment must be made in the form of a check, money order, Visa or MasterCard. Checks must be drawn on a U.S. Bank and made payable to South Bay Technology, Inc. Credit card orders by FAX may be sent to South Bay Technology at 714-492-1499. Full refund available up to January 12, 50% refund thereafter. No refunds after January 30, 1996.
In response to the microphotography manuals here are references from my EnDNote library, that I use for workshop teaching and lectures. Those from Olympus are cheap and probably free from your local Rep. or main office (1-800-446-5967 ask for Ms. Kelly who in the past send me several free samples).
(Kodak, 1980; Kodak, 1981c; Kodak, 1981a; Kodak, 1981b) (Abramowitz, 1985; Abramowitz, 1987; Abramowitz, 1990; Abramowitz, 1993) (Olympus, 1994c; Olympus, 1994b; Olympus, 1994a; Olympus, 1994d) (Inoue, 1987). ================================================================== I am not associated with the company that makes the product(s)! The information provided only constitute a professional cortesy! ================================================================== ***********Cited References:
Abramowitz M. (1985). Microscope Basics and Beyond. ed.) New York: Olympus Co.,
Abramowitz M. (1987). Contrast methods in microscopy. Transmitted light. ed.) New York: Olympus Co.,
Abramowitz M. (1990). Reflected light microscopy. An Overview. ed.) New York: Olympus Co.,
Abramowitz M. (1993). Fluorescence Microscopy. The essentials. ed.) New York: Olympus Co.,
Inoue S. (1987). Video Microscopy. ed.) New York: Plenum Press,
Kodak. (1980). Photography through the microscope. (8th ed.) NY: Eastman Kodak Company, 96 pages.
I should probably type my e-mail messages before I have my wine with dinner!
To answer the many questions I have received, the Tripod Polisher User's Group Meeting will be held during the MRS meeting in Boston as follows:
Boston Marriott at Copley Place South Bay Technology, Inc. Booth Booth #1 in the Marriott's Atrium Lounge TIME: 8:30 am Thursday November 30, 1995
I hope this clarifies things for everyone! I hope to see you there.
Best regards-
David Henriks TEL: 800-728-2233 (toll-free in USA) South Bay Technology, Inc. 714-492-2600 1120 Via Callejon FAX: 714-492-1499 San Clemente, CA 92673 USA e-mail: 73531.1344-at-compuserve.com
Would anyone know where I could get a used diffusion pump heater for an Edwards EO4 diffusion pump. I've ordered the part from Edwards but its going to be a month before I get it. It's hard to run a microprobe lab without a carbon coater. If anyone has any suggestions reply directly to me.
Thanks
Glenn ********************************************************************** * Glenn Poirier glenn_p-at-geosci.lan.mcgill.ca * * Electron Microprobe Lab Phone: (514) 398 6774 * * Earth and Planetary Sciences Fax: (514) 398 4680 * * McGill University THERE ARE THREE SIDES TO EVERY STORY; * * Montreal, Quebec YOUR SIDE MY SIDE AND THE TRUTH * **********************************************************************
I read the recent posts on TIFF and have two comments regarding Mac file formats and the Mac FTP client "Fetch". The bottom line is that Fetch by default stores Mac files in a "wrapped" format, either MacBinary or AppleSingle. These files are not TIFF files, but they contain TIFF files. To avoid this, specify "Raw binary" format when sending files using Fetch. If you specify "Ascii" format, then pixels with value 13 get changed to 10 and vice versa.
} A file either is or is not a TIFF file. If it is, then a } properly-written reader can read either the so-called MAC format } or the equally vaguely named PC format (the first two bytes of the } TIFF file define which order the bytes of multi-byte data are stored } in, and the reader has to read and act on these). I'm not an expert } MAC user, but I believe the problem with copying files to and from MACs } is that MACs add extra information to files to indicate the source } software or the type of file. These files are not, by definition, } TIFF files! There is nothing wrong with a program doing this, but it } shouldn't claim that the result is something that it is not! This seems } to me to be the source of the confusion over TIFF formats. (Perhaps a } MAC user could comment here?)
The "Mac" vs. "PC" format code in a TIFF file actually specifies how multibyte integers are stored: either least significant byte first ("little-endian", DEC PDP/11 format), or most significant byte first ("big-endian", IBM System/360 format). One might guess that the distinction arose when DEC decided to be different from IBM in the 1960's. The PC format descends from the DEC format and the Mac format descends from the IBM mainframe format. (Or maybe it arose from the right-to-left vs. left-to-right writing order of Arabs vs. Romans. When adding, we write numbers right to left...).
The TIFF spec permits integers to be stored in either format, but the same format must be used consistently throughout the file. Conforming TIFF readers are required to support both integer formats.
The important problem arises when Mac files are stored on PC or Unix file systems for the purpose of retrieval to and complete regeneration on the final destination, another Mac. This can happen either on purpose, or inadvertently, and it has nothing to do with the byte order of integers.
When stored on a Mac file system, files consist of three parts: 8 bytes of "creator code" and "file type", a data fork (which is the entire file on PC or Unix file systems), and a resource fork which would be empty for TIFF files. The creator code and file type are not essential unless you want to double click the file, all they do is determine what icon will be displayed for the file on the desktop, and which application will be launched to open the file if you double click it. If the 8 bytes are all '?', the file will be shown using a generic document icon and double clicking it will give an alert telling you there is no application to open it.
When Fetch retrieves a file from a PC or Unix file system, and you specify that the file is a Raw binary file, it will give you a chance to specify the 8 bytes of creator code and file type if you know what you want. For instance, you can make the creator code "Imag" and the file type "TIFF" and then the NIH Image TIFF icon will be shown and double clicking the file will start a copy of NIH Image if you have one. If you have more than one, it will start whichever version it feels like.
If you use Fetch and do not specify raw binary mode when sending it to a Unix or PC file system, then Fetch will pack all of the parts of the Mac file into either "MacBinary" format or "Apple Single" format. This permits another Mac to retrieve the file from the Unix or PC file system and regenerate all three parts of the Mac file in its file system. This is not what you want if you want to use the file on the Unix or PC system. One symptom of this happening is that the file gets larger by 128 bytes.
If you wish to send a Mac TIFF file to a Unix or PC system for use on that system, be sure to specify "Raw" format when using Fetch. Otherwise, Fetch will not create a TIFF file on the remote file system. It will create a MacBinary or AppleSingle file that happens to contain a TIFF file. Your Unix or PC program will then try to read the MacBinary file and say (surprise!) this is not a TIFF file.
I am looking for a used Kevex x-ray detector (any condition) late 70's and up with a +35 degree take-off angle for parts. I need the ball shaped fitting that goes between the LN2 dewar and the detector assembly but will gladly accept a whole unit.
If you can help please e-mail or call: (860) 486-2914
Thank you.
Jim Romanow Electron Microscopy Facility Physiology and Neurobiogy Department The University Of Connecticut Storrs bsgphy3-at-uconnvm.uconn.edu
WHAT: The winter meeting of the Northern California Society for Microscopy.
WHEN: Thursday, Dec. 7 from 5:30 to 9:30 p.m.
WHERE: Casablanca Banquet Facility, 979 San Pablo Ave., Albany, CA
WHO: Dr. John Mardinly Intel, Corp.
"Uses of Microscopy in the Microelectronics Industry" & Dr. Richard Demaree and Rick Giberson Cal State U. at Chico Ted Pella, Inc
"Microwave Tissue Processing for Electron Microscopy"
The Northern California Society for Microscopy (NCSM) welcomes new members. Anyone interested in microscopy of any type, from atomic force to optical will benefit from joining the NCSM. We meet four times a year to have dinner and hear about new developments in microscopy. Dues are very reasonable at only $15 a year for regular members, and $8 for students. If you are interested in joining, or knowing more about NCSM, please contact Kent McDonald by phone at 510-642-2085 during normal business hours, or by email at: klm-at-uclink4.berkeley.edu If you are interested in attending the Dec. 7 meeting described above, please call 510-486-4088 no later than Dec. 1 so we can reserve the appropriate number of dinners. Complimentary drinks will be provided by Ted Pella, Inc. and Philips Electronic Instruments and the dinner will be $16 for members, $8 for students. Please join us.
You can contact Kevex directly at 415-591-3600. They still support = their full line of detectors so this should be no problem. Since you = will need to break vacuum to change the mount, it may be best to send = the detector to Kevex and they can change the mount and ensure correct = performance before returning it to you.
----------
Fall Cleaning?
I am looking for a used Kevex x-ray detector (any condition) late 70's = and up with a +35 degree take-off angle for parts. I need the ball shaped fitting that goes between the LN2 dewar and the detector assembly but = will gladly accept a whole unit.
If you can help please e-mail or call: (860) 486-2914
Thank you.
Jim Romanow Electron Microscopy Facility Physiology and Neurobiogy Department The University Of Connecticut Storrs bsgphy3-at-uconnvm.uconn.edu
Several years ago the Microscopical Society of Canada published a book by Stanley Klosevych - "Principles and Practice of Microscopy and Scientific Photography". Information on cost can be obtained from our Executive Secretary, Marie Colbert, by email to colbertm-at-fhs.mcmaster.ca I had noted several recent postings on references to photmicrography resources. Carolyn Emerson, Editor, MSC Bulletin
An associate with a microscope collection is offering for sale a Jamin-Lebedeff set for a Zeiss Photomicroscope (fixed tube length) or other Zeiss scope with similar nosepiece and condenser mounts.
If interested, please contact Mort (Abramowitz) directly at (516) 844-5049 between the hours of 9:30AM and 3:00PM (Eastern Daylight Time) on business days.
Dr. Goldstein will begin her presentation by giving us a summary of what is happening in the Microscopy Society of America (MSA) at the national level. Following this she will give a 30 to 40 minute presentation on imaging muscle. Power spectra of electron micrographs show that the Z band lattice in mammalian muscle has at least two structural states related to the contractile state of the muscle. Cross-sectional projections of relaxed muscle show the small square pattern, while projections of fixed contracted muscle show a pattern termed the basket weave. Differing three-dimensional models constructed to visualize the transition were consistent with the cross-sectional data; therefore, we produced three-dimensional reconstruction's of Z band lattices from electron tomography of tilted specimens. The relaxed lattice shows features not imaged in micrographs of cross or longitudinal sections of muscle. Generating and analyzing these data lead to challenges in display of these "micro" structures not found in our "macro" world. Issues of perception and vision are of demonstrated importance in the display process.
PLEASE MAKE RESERVATIONS BY November 28: We need to give a "final" head count to the Cherrywood Room.
Dinner and Social Hour: $10.00 per person, $7.50 for students, payable at the door. Presentation is free for those who come later for the talk only. To reserve, contact Gib Ahlstrand at (612)625-8249, 625-9728FAX, or: giba-at-puccini.crl.umn.edu.
Social hour, dinner and the presentation will all be held in the Cherrywood Room, second floor level of the St. Paul Campus Student Center of the University of Minnesota. This location will be familiar to some who have attended our meetings there before. Parking is available nearby.
--
Gib Ahlstrand, MMS Newsletter Editor Electron Optical Facility, University of Minnesota, Dept. Plant Pathology 495 Borlaug Hall, St. Paul, MN 55108 (612)625-8249 612-625-9728 FAX, giba-at-puccini.crl.umn.edu
"MICROSCOPY & MICROANALYSIS 96" in Minneapolis, Minnesota, Aug.11-15, 1996
My daughter is involved in planning a science fair project in which she will be assessing various environmental conditions on growth of protozoan populations. What techniques are used for fixing, staining, or otherwise preparing these little guys for light microscope examination. If staining is possible to allow for segmentation from background, I could introduce her to computer image analysis. Suggestions and/or references on this topic would be greatly appreciated.
Hello, I am interested if anybody knows of any methods of visualizing cadmium in tissues treated with cadmium for the light microscope or the TEM. At the moment I have a method that employs benzothiazolylazonapthol derivatives(BTAP's) synthesized by Sumi et al. (1979, 1980, 1982) which allows for the visualization of cadmium in cadmium treated tissues for the light microscope. Unfortunately the aforesaid papers do not include the staining protocol for the use of BTAP's. So, if anybody has any experience visualizing cadmium treated tissues or knows of the protocol for the use of BTAP's or has any alternatives\suggestions, I would very much appreciate your feedback. Thank-you very much. MSCROGGIE-at-TRENTU.CA
I`m looking for a soft wich will allow me to read the *.vct file of the image from a XL 20 SEM. The image is in TIFF format wich is readable but the parameters are stored under a Philips format wich I can't read with any of my softs.
Any help is welcome
______________________________________ SCHMUTZ Marc IGBMC BP 163 F67404 ILLKIRCH Cedex France Phone 33 88 65 33 32 Fax 33 88 65 32 01 email schmutzm-at-lear.u-strasbg.fr
} I`m looking for a soft wich will allow me to read the *.vct file of the image } from a XL 20 SEM. The image is in TIFF format wich is readable but the } parameters are stored under a Philips format wich I can't read with any of } my softs. }
So am I. And I would also be very happy if anyone can give me info on software that can convert the *.img files of Philips to TIFF so that I can use the high definition images on other computers than the microscope's. We have an XL30 but that shouldn't make any difference.
Bunton Instrument Company, Inc. of Rockville, MD does this. (301) 762-5115
At 12:48 PM 11/23/95 -0500, Cannon9965-at-aol.com wrote: } Is there anyone who knows an outfit that modifies and/or builds special } custom microscopy equipment? Name and telephone if possible? } } M. Davis } } }
I now a company in France who has developped a utility for automatic decode of XL images data and transfer into a Microsoft WORD 6 document. They also have I think a converter from IMG to TIFF and a high resolution frame grabber for XL microscopes (get 3000 lines in photo mode instead of 484)
references:
I.C.I. sarl
tel (33) 84 58 02 43 fax (33) 84 54 03 98
they have no Email address but we can transfer them messages if needed.
Gatan builds accessory microscopy equipment. Don't know exactly what you want. Gatan 6678 Owens Drive Pleasanton, CA 94566 #162##193#5/463-0200 FAX 415/463-0204 _______________________________________________________________________________
Is there anyone who knows an outfit that modifies and/or builds special custom microscopy equipment? Name and telephone if possible?
M. Davis
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In the past few weeks Don Steele and Ciara Mullan have questioned the electrolytic preparation of Al and FeAl for TEM specimens. I would like to share some insight. I have found that both of these materials are conducive to the twin-jet electropolishing technique provided that the proper electrolyte and polishing conditions are applied.
For Al and some Al alloys I have found success with either 20% perchloric and 80% ethanol at -30 degress C and about 35 VDC or 20 to 25% Nitric in Methanol (add liquid nitrogen to the electroyte until it begins to solidify then begin polishing as soon as it re-liquidifies).
For FeAl either 33% nitric in methanol at -40 to -50 degrees C or 5% perchloric, 35% Butoxyth, 60% Methanol at -20 degress C and 15 VDC.
When polishing any Al alloy, it is essential that the specimen is removed immediately from the electropolisher upon perforation and thoroughly rinsed in ethanol to minimize oxidation.
I hope that this information is helpful. If there are any furbether questions, please contact me directly. Also, I will be exhibiting at the MRS meeting in Boston next week and would be happy to address any specimen preparation questions there.
Best regards,
Paul Fischione E.A. Fischione Instruments, Inc. 9003 Corporate Circle Export, PA 15632 Phone 412-325-5444 FAX 412-325-5443 e-mail paul.fischione-at-internetmci.com
Dear Group - With this note I would like to advise that we are in the process of developing a proper, comprehensive "buyers guide" on the Web for microscopy-related equipment, materials, supplies and services. In addition to links to other home pages, the system will provide source information down to the local, international level. The major challenge is to develop an index which will allow the convenient selection of ANY microscopy product or service. To this end, we would appreciate input from any interested party. From manufacturers and suppliers, we would specifically appreciate a listing of ALL categories under which they provide equipment, materials, supplies, and/or services. To them, we will provide drafts of the index as it is developed. Incidently, we do not plan on "charging" you for basic listings - but will ask for a MODEST yearly fee for extended listings. Please contact me directly, not on the listserver, regarding this matter. Regards to all, Don Grimes, Microscopy Today
Scanning Microscopy 1996 meeting May 11-16, 1996, Bethesda, Maryland (suburb of Washington, DC)
A symposium on: Scanning Probe Microscopies and Related Techniques for the Biological and Materials Sciences
will be held during the Scanning Microscopy 1996 meeting at the Hyatt Regency Hotel, Bethesda, [tutorials on May 11 and 12; scientific programs from May 13 (8:30 AM) till May 16 (noon)] The symposium organizers are:
Dr. David P. Allison, Oak Ridge National Lab., P.O.Box 2008 - M.S. 6123, Oak Ridge, TN 37831-6123, USA. (phone: 615 574 6215 / FAX: 615 574 6210 / E.mail: allisondp-at-ornl.gov);
Prof. Chunli Bai, Institute of Chemistry, Chinese Academy of Sciences, Beijing, 100080, China (Phone: 86 10 256 8158 / FAX: 86 10 255 7908 / E.mail: clbai-at-bepc2.ihep.ac.cn);
Prof. Lawrence A. Bottomley, Georgia Inst. Tech., Chemistry & Biochemistry Dept., Atlanta, GA 30332-0400 USA (Phone: 404 894 4014 / FAX: 404 894 7452 / E.mail: lawrence.bottomley-at-chemistry.gatech.edu);
Dr. Heinrich J.K. Hoerber, European Molec. Biol. Lab., Meyerhofstr. 1, D-69117 Heidelberg, Germany (phone: 49 6221 387569 / FAX: 49 6221 387306 / E.mail: horber-at-embl-heidelberg.de);
Prof. Douglas J. Thomson, Univ. Manitoba, Dept. Electrical & Computer Engg., Winnipeg MN, R3T 2N2, Canada (phone: 204 474 8797 / FAX: 204 261 4639 / E.mail: thomsom-at-ee.umanitoba.ca).
Following up on the past SPM meetings, this symposium will provide a nice occasion to present novel discoveries as well as reviews of recent developments in theory, instrumentation, and applications of scanning tunneling microscopy and related techniques, including atomic force microscopy, magnetic force microscopy, near field optical microscopy, etc. Applications of STM and other scanning probe techniques should emphasize the studies of adsorbates as well as physical and chemical process at solid surfaces. Topics of interest include: studies of processes on metal, semiconductor, and other solid surfaces: imaging of molecules, especially biomolecules; imaging of cells and other biological structures; tip-induced effects; etc. This symposium will be held simultaneously with a number of other symposia on Fundamental Physics in Microscopy and Microanalysis, Pattern Formation and Nanoscaled Structures in Thin Film Formation, Scanning Microscopy and Semiconductors: Metrology and Diagnostics (dealing with fundamental aspects of material and defect properties, characterization of surfaces and interfaces in semiconductors, properties of thin dielectrics, correlation between electrical and structural properties, new developments in characterization and imaging techniques); and biological (including microanalysis and imaging, immunolabelling, radiation effects, dentistry, corrosion casting, inner ear, bone biology, stones and crystals, etc.); Cells and Materials; and Food Structure related topics.
For the STM program, all relevant contributions are welcome with the understanding that a paper presented at this meeting must be submitted for publication in the quarterly journal Scanning Microscopy (Instructions for Authors available on request). Potential contributors are invited to discuss their ideas with one of the organizers. Suggestions for potential contributors, and participation in the program are invited. For each contribution, a completed Letter of Intent form (see over; this form can also be used to request additional information including a registra- tion/hotel form, travel support guidelines etc.) should be submitted as soon as possible (preferably to arrive at Scanning Microscopy office by December 15, 1995).
The registration fee for the entire 6-day conference (without subscription to any of the SMI journals' 1996 volumes) is either $100 (if payment reaches SMI before March 31, 1996) or $120 (from April 1, 1996); the fees with 1996 subscription to one, two or all three SMI journals (Scanning Microscopy, Cells and Materials, and Food Structure) are respectively: $180, $220, or $270 (for US delivery addresses), or $210, $270, or $340 (for outside US addresses). The room rates (exclusive of applicable taxes, currently 15%) at the Hyatt Regency Bethesda (One Bethesda Metro, Bethesda, MD 20814, USA; phone: 301 657 1234; FAX 301 657 6453) are: Single (1 person - 1 bed): $100; Double (2 persons - 1 bed) or Twin (2 persons - 2 beds): $110. Please make room reservations directly with the hotel.
Scanning Microscopy International will also be sponsoring a separate international meeting: 15th Pfefferkorn Conference on Electron Image and Signal Processing (immediately after the Bethesda Meeting) from May 18-23, 1996 at Albany, New York. The organizers are: Drs. Peter W. Hawkes (CNRS, Toulouse, France; FAX: 33-62-257999; e.mail: hawkes-at-cict.fr; as its chief organizer), W. Owen Saxton (Univ. Cambridge, U.K.) and Joachim Frank (NY State Dept. Health, Albany, NY). Possible contributors should contact one of the organizers.
For more information about the programs and publications of Scanning Microscopy, please contact Dr. Om Johari at Scanning Microscopy International.
----------- Scanning Microscopy ISSN: 0891-7035 Volume 9, Number 3, September 1995, Pages 619-926
Table of Contents
Editorial Board Inside Front Cover
Interactions of Low-Energy Electrons With Atomic and Molecular Solids (Review paper) L. Sanche 619
Work Function Dependence of Charge Transfer in Desorption and Sputtering of Atoms from Surfaces S. Steuber, P. Nordlander, H. Shao, D.C. Langreth 657
Imaging of Cherenkov and Transition Radiation from Thin Films and Particles N. Yamamoto, A. Toda 669
A Reflection Upon the Applicability of Electron Beam Induced Current (EBIC) as a Sensitive Microanalytical Technique (ppb range) for Silicon Materials Research (Tutorial paper) M. Kittler, W. Seifert 677
Photon Emission Induced by the Scanning Tunneling Microscope R. Berndt 687
Atomic Force Microscopy of Neuron Networks A. Cricenti, G. De Stasio, R. Generosi, P. Perfetti, M.T. Ciotti, D. Mercanti 695
Thermal Stability of Polystyrene Latex Self-Assembled Arrays Studied by Atomic Force Microscopy I. Nevernov, C. Nicolini 701
Atomic Force Microscopy of Nucleoprotein Complexes (Review paper) Y.L. Lyubchenko, B.L. Jacobs, S.M. Lindsay, A. Stasiak 705
Scanning Force Microscopy of Chromatin (Review paper) W. Fritzsche, J. Vesenka, E. Henderson 729
Consistency in Calibrated Backscattered Electron Images of Calcified Tissues and Minerals Analyzed in Multiple Imaging Sessions E.G. Vajda, J.G. Skedros, R.D. Bloebaum 741
Cross-Linking of Cell Surface Receptors as a Trigger of Cell Apoptosis and Proliferation Y.N. Korystov 757
X-Irradiation-Induced Disorganization of Cytoskeletal Filaments and Cell Contacts in HT29 Cells Z. Somosy, M. Sass, G. Bognar, J. Kovacs, G.J. Koteles 763
Use of Colloidal Gold and Neutron Activation in Correlative Microscopic Labeling and Label Quantitation B.J. Darien, P.A. Sims, K.T. Kruse-Elliott, T.S. Homan, R.J. Cashwell, A.J. Cooley, R.M. Albrecht 773
Confocal Laser Scanning Microscopic Studies on Alveolar Bone Remodeling with Orthodontic Tooth Movement and Retention H. Yagishita, S. Iwatsubo, T. Aoba 781
Endoarticular Loose Bodies and Calcifications of the Disk of the Temporomandibular Joint: Morphological Features and Chemical Composition C. Piacentini, C. Marchetti, A. Callegari, M. Setti, G. Bernasconi, U. Baciliero, P. Menghini, C. Brusotti 789
Scanning Electron Microscopy and Energy Dispersive Spectroscopy Analysis of Calciotraumatic Lines in Rat Labial Dentin after Acute Exposure to Strontium Chloride H. Mishima, T. Sakae, Y. Kozawa 797
The Interaction of Laser Energy with Ureter Tissues in a Long Term Investigation U. Stratmann, K. Schaarschmidt, R.R. Lehmann, A. Heinze, G.H. Willital, E. Unsold 805
In Situ Hybridization and Monoclonal Antibody Analysis of Plasma Membrane Ca-Pump mRNA and Protein in Submandibular Glands of Rabbit, Rat and Man J.L. Borke, A.E. Zaki, D.R. Eisenmann, S.H. Ashrafi, M.M. Sharawy, S.S. Rahman 817
Comparative Analysis of Patch Lesions in the Chick Inner Ear Following Acoustic Trauma: Optical Versus Scanning Electron Microscopy H.J. Adler, J. Mantooth, Y. Raphael 825
Comparison of DNA Fragmentation and Color Thresholding for Objective Quantitation of Apoptotic Cells D. Plymale, D.S. Ng Tang, C.D. Fermin, D.E. Lewis, D.S. Martin, R.F. Garry 833
Improved Methods for Preserving Macromolecular Structures and Visualizing Them by Fluorescence and Scanning Electron Microscopy (Review paper) P.B. Bell, Jr., B. Safiejko-Mroczka 843
Elicitor-Induced Resistance in Tomato Plants Against Fungal Pathogens: Ultrastructure and Cytochemistry of the Induced Response (Review paper) N. Benhamou 861
Low Temperature Techniques as a Tool in Plant Pathology (Review paper) S. Hippe-Sanwald 881
Impact of Escherichia coli on Urine Citrate and Urease-Induced Crystallization A. Edin-Liljegren, H.H. Hedelin, L. Grenabo, S. Pettersson 901
Backscattered Electron Imaging and Energy-Dispersive X-Ray Microanalysis Studies of Evidence for Calcium Salt Heterogeneity in Fifteen Gallstones from an Elderly Human (Review paper) T. Kodaka, R. Mori, K. Debari, R. Takiguchi, S. Higashi 907
List of Reviewers for Scanning Microscopy Volume 9, number 3, 1995 925-926 Scanning Microscopy International Post Office Box 66507, Chicago (A.M.F. O'Hare), IL 60666-0507, U.S.A. Telephone: (708) 529-6677 / FAX: (708) 980-6698 E.mail: 73211.647-at-compuserve.com
Form below must be completed and returned to remain / get on SMI mailing lists
Scanning Microscopy Meetings and Publications
Scanning Microscopy, Cells and Materials, and Food Structure 1996 meetings will be from May 11 to 16 at the Hyatt Regency Hotel, Bethesda, MD (suburb of Washington, D.C.). Programs on the follow- ing topics are already being planned (*fliers available, list below): *Scanning Probe Microscopies and Related Techniques for the Biological and Materials Sciences; Physical sciences programs in: *Fundamental Physics in Microscopy and Microanalysis, *Pattern Formation and Nanoscaled Structures in Thin Film Formation, *Scanning Microscopy and Semiconductors: Metrology and Diagnostics; Biological programs in: Microanalysis and Imaging, *Im- munolabelling, *Radiation Effects, Apoptosis, *Dentistry, Corrosion Casting, Inner Ear, *Bone Biology, *Stones and Crystals, etc.); *Cells and Materials; and Food Structure related topics. For participation and contribution, please complete and return the portion below. Over 400 papers were presented at the Scanning Microscopy, Cells and Materials, and Food Structure 1995 meeting from May 6-11, 1995 at Houston. Papers from these programs are being published in our appropriate journals (Scanning Microscopy / Cells and Materials / Food Structure) as processed through the review process.
Papers for publication only in our journals Scanning Microscopy, Cells and Materials, and Food Structure are invited and can be offered any time per our Instructions for Authors (printed in each journal; also, available on request, see below)
Scanning Microscopy International will also be sponsoring a separate international meeting: 15th Pfefferkorn Conference on Electron Image and Signal Processing (immediately after the Bethesda Meeting) from May 18-23, 1996 at Albany, New York. The organizers are: Drs. Peter W. Hawkes (CNRS, Toulouse, France; FAX: 33-62-257999; E.mail: hawkes-at-cict.fr; as its chief organizer), W. Owen Saxton (Univ. Cambridge, U.K.) and Joachim Frank (NY State Dept. Health, Albany, NY). Possible contributors should contact one of the organizers. Special Offers on publications: The following special discounts (expiring Dec. 31, 1995) are applicable to all listed publications (see our list over) except Scanning Microscopy and its Supplements for 1992-1996; Food Structure vol. 11-14 (1992-1996) and Cells and Materials, vol. 2-5 (1992-1996): for orders totaling $100-$299: Discount 20%; for orders totaling $300-$499: Discount 35%; and for order totaling more than $500: Discount 50%. See below for separate discounts on SM Supplements:
Orderers of Scanning Microscopy Supplement 6, 1992 "Signal and Image Processing in Microscopy and Microanalysis" may obtain Scanning Microscopy Supplement 2, 1988 on "Image and Signal Processing in Electron Microscopy" at $20 (for US delivery) or $25 (outside US delivery by sea mail) additional.
Orderers of Scanning Microscopy Supplement 7, 1993 "Physics of Generation and Detection of Signals Used for Microcharacterization" may obtain Scanning Microscopy Supplement 4, 1990 on "Fundamental Electron and Ion Beam Interactions with Solids for Microscopy, Microanalysis and Microlithography" at $20 (for US delivery) or $25 (outside US delivery by sea mail) additional.
Orderers of Scanning Microscopy Supplement 8, 1994 "Science of Biological Microanalysis" (to be released in late May) may obtain three "derived" publications [1. X-Ray Microprobe Analysis of Chemical Elements in Biology, 1993 (28 papers, 308 pp); 2. The State of Water in the Cell, 1988 (10 papers, 113 pp); and 3. Basic Methods in Biological X-ray Microanalysis, 1983 (20 papers, 284 pp)] at a special package price of $100 (for US delivery) or $115 (outside US delivery by sea mail).
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__ I wish to present at the Scanning Microscopy 1996 meeting (tentative title and summary on a separate sheet), please send a Letter of Intent form.
__ I cannot present, but am likely to attend the 1996 meeting, please keep me informed and send me: 1996 Registration / Hotel form.
__ I can neither present nor attend; please add/keep my name on your mailing list. Send me a mailing list form
Please send: 1996 program fliers (*list programs here): _________________________________________________________________ _________________________
Instructions for Authors / Major subject index / Table of Contents for: : Scanning Microscopy / Cells and Materials / Food Structure;
Flier on (see titles above and over) Scanning Microscopy: Supplement 6, 1992; Supplement 7, 1993; Supplement 8, 1994
Flier on Pfefferkorn Conferences: 1995 on Science of Biological Specimen Preparation (now over); 1996 on Electron Image and Signal Processing
Does anyone have the correct contact tel./fax numbers for the Sixth Asia-Pacific Conference on Electron Microscopy, Hong Kong, 1/5 July 1996? The two sets of numbers that I have seem to be wrong - tel.: 852-609-6845 or 852-559-9973, fax: 852-603-5031 or 852-547-9528. Thanks for your help! Regards, Don Grimes, Microscopy Today
could anybody point me at some conferences taking place in 1996, preferentially 2nd half of the year, dealing with applications of Scanning Probe techniques in the field of microelectronics (not exclusively, but there should be at least one or two sessions in this field)?
I would like information and opinions on dry nitrogen units for x-ray analysis systems. Within the next 6 months we will be installing a new SEM/FEG and we would like to replace our old dewar system. I have received some comments on the dry system; however, I would like responses from people who have had actual experience with them and can tell me the good and bad (if any) of them and how they compare with the liquid nitrogen dewars. Thank you for your help. Cathy Kelloes
Hola, Yo he recibido un mensaje de alguien de Chile que queria cambiar conocimientos sobre observaciones de ferritina en peces antarticos, pero perdi el mensaje. Si esta persona lee este comunicado y quiere cambiar ideas, estoy mui dispuesto a eso y tambien tengo interes.
Gracias y perdoneme por haber perdido la direccion. Francisco fjhblazq-at-spider.usp.br fjhblasq-at-biomed.icb2.usp.br _____________________________________________________________________________ Francisco Javier Hernandez Blazquez * Av. Prof. Lineu Prestes, 1524 Departamento de Histologia e Embriologia * 05508-900 Sao Paulo Instituto de Ciencias Biomedicas * e-mail fjhblazq-at-spider.usp.br Universidade de Sao Paulo * fjhblasq-at-biomed.icb2.usp.br ______________________________________________________________________________
I am looking for information on image analysis programs for the PC. We are in the process of implementing digital image capture and storage in the SEM lab. We have a Kevex system with the Feature Analysis and Program Automated Image analysis programs, but would like to explore using other image analysis software on the PC (486-66). The majority of our image analysis work involves counting and sizing metallic particles or feature analysis on porosity of non-biological specimens.
The ideal software would be more user friendly and versatile than the Kevex software and hopefully be low cost. I have NIH Image on my Mac, but would like to get something for the PC as well.
Any opinions and user experiences would be greatly appreciated.
You can reply to me privately (jegiles-at-space.honeywell.com) and I will post a summary of the replies.
Thanks,
John Giles Senior Materials Engineer Honeywell Space Systems
We are a Small Disadvantaged Business called Biotex Engineering and Scientific Services Inc., and we're working on a project for the Navy to determine if a better system for managing images from electron and optical microscopes can be developed. To accomplish this, we're contacting various users to determine how they are presently managing images and related data. Our goal is to develop a system that will provide fast, efficient, and affordable total image management. This is a Small Buisness Innovative Research project and any product developed will be available licenses free to Government users. Please take the time to provide the following information which will help ensure the success of our project. Thanks!
Name _____________________________Company____________________________ Loc._______________________________Phone Number_______________________
Would you have an interest in the results of this project? Yes No
What SEMs and Optical microscopes do you have?
How do you presentlly store images? (Film) (Analog: Video Laser Disc) (Digital: CD-R, Tape, HD)
If using film, would you like to see it eliminated if you could get the same quality electronically? Yes No If no, why not?
If electronic storage is used, what is the image resolution? (examp: 512 X 512)
What is the flip time (1, 2, 3 sec. or longer) from one image to another and is it too slow for your needs?) Time: Yes No
Do you network images? Yes No Like to? Yes No
What software applications do you use in image processing and management?
Can your merge and print text and images for reports? Yes No
What improvements would you like to see made in your image management capabilities?
Does Anyone in the Microscope Community have any information on the Death of Marcelle Gillott director of the EM lab at the University of Wisconson Milwaukee??? She got her Ph.D. here at Rutgers from this lab There is a Funeral service on Staten Island today and we have no idea where or when. Please send the information ASAP she was a great person, loved dearly, and we here at Rutgers need to pay our respects
If anyone in the Microscope community wishes to send flowers or condolences for Marcelle and her family her viewing will be at Martin Hughs Funeral Home, 998 Bay Street, Statin Island, New York, (718) 447-0873. There will be a service tomorrow But I'm not sure where yet.
On the PC of the XL microscope a DOS utility called MAKTIF.EXE should be present. With this utility you can convert img files (also high def) to tiff files.
There is no utility to read the vector information from a XL tiff file to a *.vct file for later use by the microscope. The vector information is stored as ASCII text under a private tag in the tiff file. So you should be able to read that part of the file as simple ascii text.
If you explain to me in more detail what you mean by "reading" the vector data, I might be able to help you out a little bit further.
We had reasonably good success isolating a SEM from building traffic vibrations by setting it on a platform that was suported by four inflated inner tubes from garden tractor tires. You could probably work out a similar scheme for a light microscope for very little cost and effort, using inflatable cushions or smaller inner tubes. Our problem, over the long run, was that after about six months the inner tubes would begin to leak, and it was a difficult matter to keep replacing them because the SEM was so heavy and unwieldy. For a light microscope, which is smaller and lighter, this should not be such a problem, and so this system might work out quite well for you. Good luck, W. C. Bigelow (bigelow-at-umich.edu)
Autoquant (518) 276-2138 sells sheets of 1/2" plexiglass with "sorbothane" feet (which they say can be stacked for even better isolation). $200 for a single 24x24" square. I'm curious how well they work. Also curious how to do a reasonable comparison of different products: how do you quantitate the effects of vibration in this application? What's sorbothane, & can I find it elsewhere?
Richard Thrift
} } } Richard Lee {richard_lee-at-QMGATE.ANL.GOV} 11/28/95 09:16am } } } Subject: Time: 11:12 AM OFFICE MEMO Vibration isolation Date: 11/28/95
Any suggestions on a quick and easy way to isolate light microscopes from vibration?
} } Any suggestions on a quick and easy way to isolate light microscopes from } vibration? } Dear Richard, Best is to absorb the vibrations, next best is to mount the LM on a platform which has no resonances near the frequencies of the vibrations. A quick method is to mount some styrofoam on a heavy material on another piece of styrofoam, or use tires or springs instead of the styrofoam. In either case, the properties of the set-up need to be matched to the prevalent frequencies. The directionality of the vibrations (if any) is also important. Good luck. Yours, Bill Tivol
} Any suggestions on a quick and easy way to isolate light microscopes from } vibration?
A slab of cement, marble etc. about 24 inches square and 2 inches thick for mass. The heavier the better. Your local paving store will oblige. Four tennis balls under the slab for air springs. OR the inner tube of a tire with the valve relocated to the outside for easy re-inflation is a more compliant isolation spring.
Next autumn we will have to get rid of a Philips EM300 (S+G) TEM to free-up some space in our lab. It still runs fine (about 700 hrs beam/year) despite its age (installed in 1972) and is under maintenance contract with Philips. It is mainly used for teaching. The main limitations are no EDS nor CBED capabilities. There are a standard stage (=893.5 =C5 resolution, without tilting capability) and a goniometer stage (=89 9=C5 resolution, =B1 60=B0 tilt).
If you are interested, please contact me and we can discuss the details of its availability (available in autumn 1996).
Thanks!
Philippe-Andre 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 // pabuffat-at-cimesg1.epfl.ch ______________________________ Eudora F2.1 ___________________________
We have to get rid of a Cambridge/Leica/LEO S-250 SEM to free-up some space in our lab. It still runs fine (=895=C5 resolution-at-20keV). It was installed in 1980 and was is under maintenance contract with Cambridge until the end of 1993.
If you are interested, please contact me and we can discuss the details of its availability (available in summer1996).
Thanks!
Philippe-Andre 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 // pabuffat-at-cimesg1.epfl.ch ______________________________ Eudora F2.1 ___________________________
Next summer we will have to get rid of a Cambridge/Leica/LEO S-100 SEM to free-up some space in our lab. It still runs fine, about 1100 hrs beam/year with students and SE/BSE observation up to =8920'000 X (=895=C5 resolution-at-20keV). It was installed in 1983, it is under maintenance contract with Cambridge and its electronics has been entirely rejuvenated (bad contacts) two years ago.
If you are interested, please contact me and we can discuss the details of its availability (available in summer1996).
Thanks!
Philippe-Andre 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 // pabuffat-at-cimesg1.epfl.ch ______________________________ Eudora F2.1 ___________________________
Try A.Q.Sorbothane feet - avaiable at 800-229-0644 for about $60.00 these set of four hockey puck style "rubber" feet should cut vibration on most scopes. if not try a Kinetica air table at $3500.00
Scott E. Berman Advanced Imaging Concepts, Inc. Princeton, NJ (908) 274-1877 x26 (908) 274-1974 Fax
Richard, you can find Sorbothane or other vibration absorbing products in any audiophile magazine - a big mail order company that would have it is Audio Advisor at 800-942-0220 4 sorbothane big feet should run ~$50.00 and for heavier scopes ther are Sims Navacom silencers for about $75.00. Or try The Needle Doctor 800-229-0644
Scott E. Berman - Audiophile and Digital Imaging Expert Advanced Imaging Concepts Princeton, NJ (908) 274-1877 x26 (908) 274-1974
As you can imagine, we have a lot of situations where high mag light microscopes are close to or mounted on rotating machinery. We've been using corrugated rubber matting very successfully. The pieces of matting are the same size as the base of the scope.
Unfortunately I don't know of sources of this material. We bought it so many years ago that that information is long gone. You might venture down to your friendly machine shop guys and look through their tool distributor catalogs (MSC, McMaster-Carr, etc.).
Good luck,
Joe Tabeling Delaware Diamond Knives 800-222-5143 http://www.ddk.com/ddk
} Hi everybody, } } Can someone please enlighten me as to the meaning of "ploem illumination"? } I have looked up several dictionaries to no avail. } } Thanks, } Felicity
Ploem illumination is one type of epi-illumination for fluorescence in which the excitation filter, dichroic mirror, and barrier filter for a specific (set of) fluorochrome(s) are mounted together in one cube. Different cubes are fitted into a revolver in the microscope stand and can be switched easily. It is the type of epi-illumination used by Leica a.o.
Another question: Felicity, I noticed that you also had posted your question to a confocal-list. I would be very glad to get subscription information on that.
} Hi everybody, } } Can someone please enlighten me as to the meaning of "ploem illumination"? } I have looked up several dictionaries to no avail. } } Thanks, } Felicity
J. S. Ploem introduced already 1967 dichromatic mirrors for incident illumination (epi-illumination) in fluorescence microscopy. The exciting radiation is directed downwards from the light sporce to the object by the dichromatic beam splitter which acts as a condenser. The fluorescent emission passes back through the objective and the dichromatic beam splitter and the barrier filter to the observer. Ploem, J.S., Z. Wiss. Mikr. 68, 129-142. Hope this will help you. Sverker
********************************************************* Sverker Enestrom M.D., Ph.D. Department of Pathology University of Linkoping, Sweden Phone: +46 13 22 15 20 Fax: +46 13 13 22 57 *********************************************************
To the Microscopy List Earlier this month I posted the Confocal vs Decon question and have been getting many responses. At the moment we the Leica system (confocal) on demo, and in Jan we will have the Scanalytics Decon system for two days. I have been collecting all the responses and will soon create a summary, together with our demo experiences. Please be patient and keep those opinions coming. Thanks to all for your interest,
Mike Delannoy Microscopy Facility JHMI Baltimore, Md
Hello, I have recently started doing TEM on the infection of sorghum leaves by the fungus Colletotrichum graminicola. However, I have not had very much success as of yet because I have had a lot of trouble fixing and sectioning the leaf tissue. My main problems at the moment are poor fixation and the resin splitting away from the leaf along the cuticle when I am sectioning.
I was wondering if anyone had done TEM on sorghum before or if anyone had any suggestions to how I could get round the problems I mentioned above.
I would be very grateful to recieve any suggestions.
P. Wharton
IACR-Long Ashton Research Station University of Bristol UK
} What's sorbothane, & can I find it elsewhere? } Dear Richard, My guess is that it is polyurethane--strictly a guess based on its property of absorbing energy. Besides it sounds logical. Yours, Bill Tivol
We are currently looking into the possibility of utilizing a Video Archival Retrieval System to store our SEM images rather than Polaroids.
The VARS that we got the sales pitch on stores the image signals on a optical memory disk recorder as the primary storage medium. The disk can store between 36k and 54k different images.
I would like to hear from anyone who has knowledge of these types of systems.
Pros/Cons/Opinions would be most helpful.
Steve McGarvey Photolithograhy Process Tech Fujitsu Microelectronics, Inc. 21015 S.E. Stark Street Gresham, Oregon, USA 97030-2099
I've seen something similar while imaging silicates with our Cameca SX50 at higher KeV (25KeV) rather than what we typically use (15KeV). I had to question the video amplifier because it didn't make sense. What I should have tried then, and what I might suggest for you to try is lowering the beam current for the more intense image signal ...
cheers, shaf
{\/} /\ {\/} /\ {\/} /\ {\/} cognito, 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/epmahome/
We are upgrading an AMRAY 1600 tungsten SEM. We will be replacing a TN5600 Be-window EDS and the electronics side of a MicroSpec WDX-2A WDS. We will be keeping the WD spectrometer itself. We would like to have one integrated
ED/WD system running on an Intel PC using Windows and appropriate instrument hardware. We have contacted a couple of vendors that say they can provide acceptable solutions and we will be running samples on their equipment soon.
Do any of you have experience with the Oxford Link/ISIS system with the WDX-2A? How about a Kevex integrated system? Any other systems?
Harold J. Crossman OSRAM SYLVANIA INC. Lighting Research Center 71 Cherry Hill Dr. Beverly, MA 01915 (508) 750-1717
Although I'm not certain whether this is the effect you're seeing, it is somewhat well-documented that electron channeling contrast will exhibit contrast reversals upon certain changes in energy. The way to check whether channeling is important here is simply by tilting your sample and seeing whether the contrast changes, for a constant beam energy. I looked at your web images, and at first glance, it wasn't apparent to me whether that was a channeling effect.
Another possibility: is there a chance that at the lower energy, you are sampling some kind of thin, high Z overlayer in the bright regions (contamination or another phase or ...) which is for the most part fully penetrated at 25 kV? For a very thin overlayer, the EDS signal may not see it well.
Regards,
Bob Keller NIST Materials Reliability Division Boulder, CO
Thanks to all who responded to my questions concerning teaching TEM courses. As promised, I am posting this summary of the responses I received (in a slightly edited fashion). In addition, I have summarized my findings in this rough table:
How often (times per year) How many students Student Fee? 1 12 No 1 12 No 1 No 0.5 - No 1 12 yes 1 5 No 1 20-30
1 3-5 No 1 5 No 0.5 8-20 No 1 7 $150 0.5 14 $50 1 10-30 No 1 8 No 1 12 No 1 10-30 No
So, I conclude most of us are teaching the course once a year for 12 students without charging any unusual fee.
} 1. How often do you teach a TEM course? Once per Fall.
} 2. How many grad/undergrads take it? About one dozen.
} 3. Do you charge for consumables or beam time? No, but typically users initiate long term microscopial investigations that do generate "U revenue."
} 4. Does it make use of a departmental or campus wide facility? If so, } does it interfere with researchers using the facility?
Departmental. Two lab sessions meet on Thursdays. I suspect a poll of current users would reveal that this constitutes minimal interference. We work every day, all hours anyway. Of course lab time is coordinated with two one hour lectures per week, and a dedicated microscopy education computer network.
Best regards,
Mark T. Biedrzycki Computer Networking Laboratory Materials Science and for Microscopy Education (CNLME) Engineering Department at the University of Arizona, Tucson
Donald L. Lovett e-mail: lovett-at-trenton.edu Asst. Professor, Dept. of Biology voice: (609) 771-2876 Trenton State College, NJ 08650-4700 fax: (609) 771-2674
The TEM course is offerred once a year. We "offer" a certain amount of free beam time to induce interest in the course. However, students are expected to complete a project, as well, for which their advisors are expected to pay for the beam time, which is "discounted" from the normal rate. The course users have priority over researchers, since the primary function of a university is to teach.
At Connecticut College, a small liberal arts college, I teach an EM (TEM and SEM) course every other year. The students are all undergraduate. There are no fees for the students; my department (i.e. me) runs the microscopes. I do work very hard on keeping the costs of consumables down, especially film. We will be adding a digital SEM soon, so I expect a lot of laser prints rather than expensive Polaroids. Course expenses come out of the academic supplies budget. I know our situation here is probably quite different compared to a large university setting.
Page Owen Connecticut College Dept. of Botany tpowe-at-conncoll.edu
Greetings from the University of Tennessee
} 1. How often do you teach a TEM course?
Once a year in the Spring semester } 2. How many grad/undergrads take it?
Typically 3-4 for the Biological course (i.e Histology) and perhaps 8-10 for the materials science/diffraction course.
} 3. Do you charge for consumables or beam time?
Yes for the Histology course, no for the Materials Science (this anomaly is the result of an historical accident).
} 4. Does it make use of a departmental or campus wide facility? If so, } does it interfere with researchers using the facility?
Yes and yes, but the first job of universities is to teach so my research and every one elses can reasonably be expected to wait once in a while.
David Joy Professor (Biochemistry, Molecular and Cellular Biology) Director EM Facility Phone/FAX (423)-974-3642
I've been teaching TEM here at Eastern, a four year undergraduate institution, for 18 years. The course is offered in the fall semester only. There are many reasons for this. The main reason is that I have other teaching responsibilities and no room in my life for two semesters of EM. The other main reason is that I like to train students in the fall so that if they really like EM, they can take an independent study under me in the spring. This also helps me in my research since I don't have any graduate students. When teaching the 4 credit EM course, I also require that all of the students take a one credit "mini" course in Biological Ultrastructure. This course lasts for 5 weeks at the beginning of the semester. It is also during those first 5 weeks that the students are undergoing 'basic training" in the EM course. Around the time the ultrastructure course is over, the students are just beginning to look at their materials under the scope and they can now actually identify all of the organelles they learned about during the previous five weeks. As far as the number of students, my maximum and minimum is 5. As you probably know, its hard to tell an administrator that you can only teach a course to 5 students, especially a couse that is this expensive! Every time we get a new dean or academic VP, I have to re-explain the course and why I can only teach 5. By the way, I've never had any trouble getting five for the course. There is usually a one to two year waiting list. As I mentioned above, the course is expensive. Not just for supplies, but also for repair and maintenance. The school has always taken care of maintenance and repair (its been a struggle on my part to keep costs down) and a very small amount of supply money. I have, on a few occasions, asked the students to buy their own film (about $50). Most of the supply money comes from my research grants (a fact that I do not tend to spread about).
We are a very small department and I am the only one using the TEM, so it is not a campus-wide facility. I just recently got an SEM and several people are interested in using it. We will have to work something out. I hope I have been some help to you. Don't hesitate to contact me if you need any further information.
Martin A. Levin Department of Biology Eastern Connecticut State University Willimantic, CT 06226 (860) 465-4324
I was involved in a TEM course when I was at Yale University. The facility I ran was a core TEM/SEM facility for the Biology and Molecular Biology/Biophysics departments. I was responsible for teaching the Lab course for the undergrad. course in Cell Biology. The lab consisted of learning basic skills in LM and TEM. I had one term of 20 to 30 students per day, five days a week, and two Graduate TAs per day, who also needed instruction. My goal was to keep the TAs at least two weeks ahead of the students. It was alot of work for me, but I really enjoyed it.
I also taught a Summer course to train Graduate students and Post Docs. I usually got from 7 - 10 individuals per year. As it was my responsibility to train and supervise anyone in the departments with a demonstrated need for microscopy, it was easiest to teach a group rather than individuals.
Both courses were extremely rewarding. There were always a few individuals who mearly wanted to get by, but there were also those with a serious love for the technique. Cheers,
Doug Keenee
At the University at Albany, State University of New York, we: 1. Teach a combined SEM/TEM class once/year. We also offer an advanced class on image processing. 2. Class size is typically very small (3-5 students). During the late 70s, early 80's the class was typically 5-10 students. With growing interest in structural biology/molecular imaging, we hope enrollment will grow to past levels. 3. No charges beyond tuition. Yet. 4. The class uses equipment at the University-wide EM facility (2 TEMs, 1 SEM), as well as those at the Wadsworth Center, NY State Dept. Health (1 antique SEM, 4 conventional TEMs, and intermediate-voltage TEM, and the 1.2MeV machine). There is plenty of beam time for all.
I'd be interested in your findings. Regards, Sam Bowser -------------------------------- Sam Bowser, Ph.D. Wadsworth Center NY State Department of Health Albany, NY 12201-0509 (518) 473-3856 bowser-at-wadsworth.ph.albany.edu
} 1. How often do you teach a TEM course? I teach it every semester if there is a demand for it by 4 or more students. I would prefer to teach it only once a year
} 2. How many grad/undergrads take it? Enrollment varies. It averages 5 students, mostly graduate students. I have been trying to train undergrad who will stay long enough to work on a research project with a professor before leaving for other endeavors. This has been hindered by the fact that tours were previously arranged by the upper division courses. Lately we have been bringing in the freshman biology class on tours and this has sparked new interest. It is too early to tell if this will produce more students at a lower grade level. Our tours involve working equipment (i.e., slicing microtomes, grids in TEM, samples in SEM, thick stained sections on LM, etc) } 3. Do you charge for consumables or beam time? We receive some class supply money and the service contract is covered by the dept. If we get trained students we can expand our researcher pool by putting students to work with professors. Researchers do pay beam time and consumables, but usually are loath to take the time to train or seek out students for EM.
} 4. Does it make use of a departmental or campus wide facility? If so, } does it interfere with researchers using the facility? Most users have been trained here in the Facility and grumble first as students about researchers time, then as researchers about student time. Generally not a problem, although this is a reflection in part of the limited number of users in the Facility. At lab report time, scope time is a premium, otherwise it is usually available on a walk in basis.
Dr. Steve Barlow EM Facility/Biology Dept. San Diego State University San Diego CA 92182-4614 phone: (619) 594-4523 fax: (619) 594-5676 email: sbarlow-at-sunstroke.sdsu.edu
Dear Tom; In response to your inquire. We (One other professor and myself) teach a course called Anatomy 660: Introduction and practice of Electron microscopy. We teach it every other summer school. It runs for 8 weeks and meets every afternoon from 1:00 to about 4:00. There is a hour to two hour lecture and then a demonstration of what subject the lecture was about. It was designed this way for the student to then start a project and work on it for the remaining 4 weeks, for a 12 week summer session total, using any of the techniques they learned about. They supply their own supplies then during that time. They can use the facility free of charge for that time. We call in about 6 other "experts" on our campus to teach special subjects. ie: Immuno, confocal, steriology, etc. We usually have from 8 to 20 students mostly graduate. one or two advanced undergrads. We use the EM Facility and the hope is to eventually attract new users for the facility. We need more customers. Please let me know what your survey shows. If you have more questions please contact me again. Thanks Grayson Grayson Scott 1300 Univ. Ave Univ. of Wisconsin Electron Microscope Facility Madison, Wisconsin 53706 608-262-2993 Fax 608-262-7306 '''
} 1. How often do you teach a TEM course? We teach one TEM course in the fall and SEM in spring and summer semesters. TEM is not as popular as SEM and is certainly more expensive to run. Its days may be numbered here. SEM is thriving, however.
} 2. How many grad/undergrads take it? We limit the TEM course to 7 and the SEM to 12. Grad students only. There is another course on campus for undergrads but this is lecture and lab/demonstration only. They come to our facility for a 2 hr demo of the EM's. Specimen prep, interpretation, etc is taught by the lecturer in the other course. Really interested students then take our course for more hands on training. This helps to weed out students who are not seriously interested.
} 3. Do you charge for consumables or beam time? $150 to take the course. Covers 10 hr scope use. Students must buy all consumables.
} 4. Does it make use of a departmental or campus wide facility? Campus wide facility
If so, does it interfere with researchers using the facility? We have two levels of scopes,older ones for students and newer ones for researchers. We may stop this in the future as it is too expensive to maintain for the benefits (cash) derived from students.
Contact me if you need more info.
############################################################################ # John J. Bozzola, Director Center for Electron Microscopy 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 ############################################################################ #
Background: "Comprehensive University" (as distinct from "Research University", although I find the appelation redundant); 5200 students 4500 of them undergrads. Master's program only, no PhD offered. Roughly 275 biology majors, 12 biology faculty. Research expected of faculty (roughly 1 paper/2 years), involvement of undergrads in faculty research expected. } } 1. How often do you teach a TEM course? Every other year, although I will train students in SEM at any point that their research project may demand it. 222. How many grad/undergrads take it? Last offering, 14 signed up; nine stuck with it. Two of the nine were grad students. } } 3. Do you charge for consumables or beam time? No. Students taking the course pay a $50 lab fee (lab fees for other courses in the dept. are $15-$25). Students must buy their own fine-point foreceps, lab manual, and photographic paper in excess of 50 sheets. We have no method for charging for beam time; external users are welcome to use the facility if they pay for plate film or polaroids (35mm is free). } } 4. Does it make use of a departmental or campus wide facility? If so, does it interfere with researchers using the facility? The facility is departmental. Given the focus on research that involves undergrads, the course not only doesn't interfere with faculty research, it enhances it. Service contracts are paid out of the departmental base budget, which was increased by the necessary amounts as the microscopes were added.
All-in-all, a pretty cool situation here. HTH Julian Smith III Biology Winthrop University
University of Illinois -at- Urbana-Champaign
I am pleased to respond to your request for information about teaching EM. It has been a dismal situation here for the past 3 years since the campus biotechnology center has taken over the operations of the Center for Electron Microscopy. Biotech refused to support instruction as a part of their policy (that is now under review and may be reversed). What happened was that I made an effort to continue some line of instruction, but I do it without TA's, as an overload assignment without extra compensation, and by taking money from my research funds to pay the CEM/Biotech. for any instructional use of equipment and facilities. The single course now offered is termed: Integrated Electron Microscopy, but has a good deal of other microscopies also included along with microanalysis. Because it was just too much for me, there is no lab, but I have fashioned the course to have weekly demonstration periods. The demos are mostly at the Center for Electron Microscopy, but include sites at other places on campus as well. All lab courses have been dropped. The only way students get instruction is to either do an independent research project with a participating faculty member (not many of them), or to pay the CEM for a staff member to give tutorial assistance. For a student with no background this may be about $1,000 to $2,000. Not many takers.
} 1. How often do you teach a TEM course?
The Integrated EM course is offered once per year--but due to my schedule may need to go on alternate year basis. (Previously, the courses were offered both semesters--though not summers--each year.)
} 2. How many grad/undergrads take it?
We have been enrolling about 20-30 students, mostly grads each time, but this semester we have only 10. One student is an undergrad.
} 3. Do you charge for consumables or beam time?
I do not request a fee from the students since we have demos only. As I indicated above, I pay for the demo time.
} 4. Does it make use of a departmental or campus wide facility?
Yes--although the facility has become more of a service lab than a multi-user facility.
If so, } does it interfere with researchers using the facility?
Not to my knowledge. There is plenty of available time on the equipment.
} If you e-mail me directly I will be happy to post a summary of the } responses to the list once they are all in.
} Thanks for your input.
You are welcome.
************************** Richard F. E. Crang Professor of Plant Biology University of Illinois (217) 244-3143 **************************
} 1. How often do you teach a TEM course? Once per Fall.
} 2. How many grad/undergrads take it? About one dozen.
} 3. Do you charge for consumables or beam time? No, but typically users initiate long term microscopial investigations that do generate "U revenue."
} 4. Does it make use of a departmental or campus wide facility? If so, } does it interfere with researchers using the facility? Departmental. Two lab sessions meet on Thursdays. I suspect a poll of current users would reveal that this constitutes minimal interference. We work every day, all hours anyway. Of course lab time is coordinated with two one hour lectures per week, and a dedicated microscopy education computer network.
Best regards,
Mark T. Biedrzycki Computer Networking Laboratory Materials Science and for Microscopy Education (CNLME) Engineering Department at the University of Arizona, Tucson
1. How often do you teach a TEM course? ==} Basic SEM and TEM undergraduate course taught in fall semester each year. Advanced TEM taught every other year in the spring semester, alternating with Advanced SEM and X-ray Microanalysis.
2. How many grad/undergrads take it? ==} 15-30 students take the undergrad SEM/TEM course, aand perhaps half of these are actually graduate students. About 10 graduate students take the Advanced TEM course and the Advanced SEM and X-ray Microanalysis course.
3. Do you charge for consumables or beam time? ==} Both are charged to the courses making them rather expensive to run since each course has approximately 10 different laboratories.
4. Does it make use of a departmental or campus-wide facility? If so, does it interfere with researchers using the facility? ==} All electron microscopes are in the campus-wide central facility located in the Dept. of Materials Science and Engineering. All departments including biology and geology use the instruments in this department. In addition to our six research electron optics instruments, we maintain three 1970s-vintage microscopes for use by our undergraduate and graduate students: a Philips EM301 TEM and two ETEC Autoscan SEMs.
CE Lyman
Charles E. Lyman Phone: (610) 758-4249 Prof. of Materials Science and Engineering Fax: (610) 758-4244 Lehigh University E-mail: cel1-at-lehigh.edu 5 East Packer Avenue Bethlehem, PA 18015-3195
Thomas E. Phillips, Ph.D. Associate Professor of Biological Sciences Director, Molecular Cytology Core Facility 3 Tucker Hall University of Missouri Columbia, MO 65211 (314)-882-4712 (voice) (314)-882-0123 (fax)
A few requests have come to post a summary of responses to a question about using EDS for quantitative analysis of C and O in steels. Here it goes ...
DETECTABILITY. Carbon content for most of steels ranges from 0.05 wt % to 0.95 wt %, and in some special cases reaches 4 wt%. Solubility of oxygen in solid iron is very poor, so that the total oxygen content, with few exceptions, is bellow 100 p.p.m. Considering the EDS detection limit for light elements is about 0.5 wt%, the quantitative analysis of both C and O can be a problem. Not less then 1% (another author gives 5%) can be quantitatively analysed by EDS.
STANDARDS. At the risk of stating the obvious ! - Reliable , accurate Quantitative Microprobe Analysis depends very much on having good homogeneous Standards of known composition . If you are generating Quantitative data on an element which is present usually at { 1 wt % in a "matrix " which is usually } 95% Iron you need a standard which resembles your unknown , what I mean is : you cannot really use a mineralogical type standard of several percent carbon as a standard for steels , ZAF effects between Standard and unknown become too marked. You really need a Steel or Range of Steels as your Standards.
MICROSTRUCTURE, MICROSEGREGATIONS, INCLUSIONS. When doing E.P.M.A. on Micron sized areas of a steels the underlying Microstructure becomes important. When probing a steel which has a coarse Pearlitic phase , your probe spot hits a Cementite lamellae in this region , your Carbon counts will skyrocket but will bear no resemblance to the true, bulk Carbon value of your sample. Area raster analysis with some beam defocussing, at several sites on your sample is one way to compensate for Microstructural effects. Much of the oxygen in steels is tied up in discrete non metallic inclusions eg.. Silicates , Aluminates . In others there are localised Oxygen "hot spots" at discrete sites throughout the steel matrix all with the potential to give misleading results !
OTHER TECHNIQUES. Because it is problematic to analyse O an C in steels with EDS, a number of other technique were suggested.
WDS has better minimum detection limits and better signal discrimination.
BE detector + image analysis. The oxygen that was dissolved in liquid steel comes out as non-metallic inclusions. These are commonly aluminates, silicates, and other oxides. So determining the content of inclusions is an image analysis problem, not one for X-ray analysis. The area fraction of the inclusions times their oxygen stoichiometry gives an answer, however, inclusions are generally determined by standard procedures such as ASTM E 45. The carbon in high carbon steels is usually in the form of iron carbides. For example, the optically identifiable phase, "pearlite" is actually a fine mixture of ferrite with cementite (Fe3C). The carbon concentration in cementite is stoichiometric at 6.7 wt.%, and need not be analysed. The area fraction of pearlite by image analysis correlates with the carbon content of the steel.
EELS can be useful for light elements.
CALCULATION OF THE DETECTION LIMIT. Macintosh computer program DTSA--available from the U.S. Natl. Inst. of Standards and Technology (NIST)-- is to calculate the detection limits and lots more. DTSA does first-principles computer simulation of EDS spectra with user-selected experimental variables (detector parameters, electron beam parameters, physics choices, etc. By simulating the spectra before doing any experimental work, you can determine the detection sensitivities, assess spectral interferences and absorption/fluorescence effects, and devise an optimum strategy for the experimental analysis.
ABSOLUTELY POSITIVE RESPONSE as to the quantitative EDS of C and O in steels came from Mark T. Biedrzycki (Computer Networking Laboratory, Materials Science and for Microscopy Education (CNLME), Engineering Department the University of Arizona, Tucson):
In general, this (EDS analysis of C and O in steels) depends on the instrument you are using and what type of detector; the short answer is pretty decent qualitative and quantitative analyses. We use Hitachi microscopes and NORAN EDS apparatus to good end routinely. If you would like recent publications detailing findings, please let us know. =====================================================
Thank you very much all contributed to the topic:
Michael Griffiths B.H.P. Rod & Bar Products Div Newcastle. griffiths.michael.mj-at-bhp.com.au
John J. Friel Princeton Gamma-Tech 1200 State Rd. Princeton, NJ 08540 jjf-at-pgt.com
Alan Stone ASTON Metallurgical Services 73004.1733-at-compuserve.com
Larry Thomas Battelle, Pacific Northwest Lab Richland, WA, USA l_thomas-at-ccmail.pnl.gov
Mark T. Biedrzycki Computer Networking Laboratory Materials Science and for Microscopy Education (CNLME) Engineering Department at the University of Arizona, Tucson mtb-at-sodium.sem.Arizona.EDU
Ubirajara P. Rodrigues-Filho Instituto de Qu=EDmica- Universidade Estadual de Campinas Campinas, SP, Brazil ubira-at-iqm.unicamp.br
Brian G. Demczyk demczyk-at-ERXINDY.rl.plh.af.mil
--- Alex Titkov Electron Microscopy Unit RMIT Applied Physics GPO Box 2476V Melbourne VIC 3001 AUSTRALIA alex-at-bunyip.ph.rmit.edu.au Voice:(03) 9660 2205 Fax: (03) 9660 3837
} Subject: Time: 11:12 AM } OFFICE MEMO Vibration isolation Date: 11/28/95 } } Any suggestions on a quick and easy way to isolate light microscopes from } vibration?
Tennis balls under a wooden frame, on which the LM sits. We have had 6 under a large Zeiss Universal for years and we are right next to a busy road on the 2nd floor. Works like a dream!
Diana van Driel Dept Ophthalmology Sydney University AUSTRALIA 2006
The Zeiss confocal has some sort of airbag/tyre system for vibration isolation, like the suggested tractor inner tubes. We've used a bicycle inner tube about 3/4 filled with water under a heavy terrazzo (fake marble) slab - seems to work OK.
____________________________________________________________ Rosemary White __ / Department of Ecology _/ \__/ \ and Evolutionary Biology / \ Monash University / Australia \ Clayton, Victoria 3168 \ ____ / phone 61-3-9905 5670 \_/ \_*_/ fax 61-3-9905 5613 __ email r.g.white-at-sci.monash.edu.au \/
We are looking to upgrade the teaching TEM in our EM Center. If you have a microscope that you are considering surplussing or trading in, please let me know.
Also, if you are interested in obtaining a basic, functional scope, let me know.
Trades will be considered.
Please contact me directly, not the microscopy list, for further details.
We had reasonably good success isolating a SEM from building traffic vibrations by setting it on a platform that was suported by four inflated inner tubes from garden tractor tires. You could probably work out a similar scheme for a light microscope for very little cost and effort, using inflatable cushions or smaller inner tubes. Our problem, over the long run, was that after about six months the inner tubes would begin to leak, and it was a difficult matter to keep replacing them because the SEM was so heavy and unwieldy. For a light microscope, which is smaller and lighter, this should not be such a problem, and so this system might work out quite well for you. Good luck, W. C. Bigelow (bigelow-at-umich.edu)
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We are currently looking into the possibility of utilizing a Video Archival Retrieval System to store our SEM images rather than Polaroids.
The VARS that we got the sales pitch on stores the image signals on a optical memory disk recorder as the primary storage medium. The disk can store between 36k and 54k different images.
I would like to hear from anyone who has knowledge of these types of systems.
Pros/Cons/Opinions would be most helpful.
Steve McGarvey Photolithograhy Process Tech Fujitsu Microelectronics, Inc. 21015 S.E. Stark Street Gresham, Oregon, USA 97030-2099
Dear Microscopists: I have a student who has prepared specimens of the Byrozoan "Cristatella mucedo" for the SEM. Unfortunately a literature search has not turned up much information on the surface morphology of this specimen or closely related ones. The references he has obtained so far offer limited light microscopy images and diagrams only. I was wondering if anyone out there knows of any references containing SEM micrographs of this organism. To save listserver bandwidth any responses can be sent to my personal E-mail address - Bray-at-HG.ULETH.CA Thanks in advance Doug Bray
Thanks to all who responded to my questions concerning teaching TEM courses. As promised, I am posting this summary of the responses I received (in a slightly edited fashion). In addition, I have summarized my findings in this rough table:
How often (times per year) How many students Student Fee? 1 12 No 1 12 No 1 No 0.5 - No 1 12 yes 1 5 No 1 20-30
1 3-5 No 1 5 No 0.5 8-20 No 1 7 $150 0.5 14 $50 1 10-30 No 1 8 No 1 12 No 1 10-30 No
So, I conclude most of us are teaching the course once a year for 12 students without charging any unusual fee.
} 1. How often do you teach a TEM course? Once per Fall.
} 2. How many grad/undergrads take it? About one dozen.
} 3. Do you charge for consumables or beam time? No, but typically users initiate long term microscopial investigations that do generate "U revenue."
} 4. Does it make use of a departmental or campus wide facility? If so, } does it interfere with researchers using the facility?
Departmental. Two lab sessions meet on Thursdays. I suspect a poll of current users would reveal that this constitutes minimal interference. We work every day, all hours anyway. Of course lab time is coordinated with two one hour lectures per week, and a dedicated microscopy education computer network.
Best regards,
Mark T. Biedrzycki Computer Networking Laboratory Materials Science and for Microscopy Education (CNLME) Engineering Department at the University of Arizona, Tucson
Donald L. Lovett e-mail: lovett-at-trenton.edu Asst. Professor, Dept. of Biology voice: (609) 771-2876 Trenton State College, NJ 08650-4700 fax: (609) 771-2674
The TEM course is offerred once a year. We "offer" a certain amount of free beam time to induce interest in the course. However, students are expected to complete a project, as well, for which their advisors are expected to pay for the beam time, which is "discounted" from the normal rate. The course users have priority over researchers, since the primary function of a university is to teach.
At Connecticut College, a small liberal arts college, I teach an EM (TEM and SEM) course every other year. The students are all undergraduate. There are no fees for the students; my department (i.e. me) runs the microscopes. I do work very hard on keeping the costs of consumables down, especially film. We will be adding a digital SEM soon, so I expect a lot of laser prints rather than expensive Polaroids. Course expenses come out of the academic supplies budget. I know our situation here is probably quite different compared to a large university setting.
Page Owen Connecticut College Dept. of Botany tpowe-at-conncoll.edu
Greetings from the University of Tennessee
} 1. How often do you teach a TEM course?
Once a year in the Spring semester } 2. How many grad/undergrads take it?
Typically 3-4 for the Biological course (i.e Histology) and perhaps 8-10 for the materials science/diffraction course.
} 3. Do you charge for consumables or beam time?
Yes for the Histology course, no for the Materials Science (this anomaly is the result of an historical accident).
} 4. Does it make use of a departmental or campus wide facility? If so, } does it interfere with researchers using the facility?
Yes and yes, but the first job of universities is to teach so my research and every one elses can reasonably be expected to wait once in a while.
David Joy Professor (Biochemistry, Molecular and Cellular Biology) Director EM Facility Phone/FAX (423)-974-3642
I've been teaching TEM here at Eastern, a four year undergraduate institution, for 18 years. The course is offered in the fall semester only. There are many reasons for this. The main reason is that I have other teaching responsibilities and no room in my life for two semesters of EM. The other main reason is that I like to train students in the fall so that if they really like EM, they can take an independent study under me in the spring. This also helps me in my research since I don't have any graduate students. When teaching the 4 credit EM course, I also require that all of the students take a one credit "mini" course in Biological Ultrastructure. This course lasts for 5 weeks at the beginning of the semester. It is also during those first 5 weeks that the students are undergoing 'basic training" in the EM course. Around the time the ultrastructure course is over, the students are just beginning to look at their materials under the scope and they can now actually identify all of the organelles they learned about during the previous five weeks. As far as the number of students, my maximum and minimum is 5. As you probably know, its hard to tell an administrator that you can only teach a course to 5 students, especially a couse that is this expensive! Every time we get a new dean or academic VP, I have to re-explain the course and why I can only teach 5. By the way, I've never had any trouble getting five for the course. There is usually a one to two year waiting list. As I mentioned above, the course is expensive. Not just for supplies, but also for repair and maintenance. The school has always taken care of maintenance and repair (its been a struggle on my part to keep costs down) and a very small amount of supply money. I have, on a few occasions, asked the students to buy their own film (about $50). Most of the supply money comes from my research grants (a fact that I do not tend to spread about).
We are a very small department and I am the only one using the TEM, so it is not a campus-wide facility. I just recently got an SEM and several people are interested in using it. We will have to work something out. I hope I have been some help to you. Don't hesitate to contact me if you need any further information.
Martin A. Levin Department of Biology Eastern Connecticut State University Willimantic, CT 06226 (860) 465-4324
I was involved in a TEM course when I was at Yale University. The facility I ran was a core TEM/SEM facility for the Biology and Molecular Biology/Biophysics departments. I was responsible for teaching the Lab course for the undergrad. course in Cell Biology. The lab consisted of learning basic skills in LM and TEM. I had one term of 20 to 30 students per day, five days a week, and two Graduate TAs per day, who also needed instruction. My goal was to keep the TAs at least two weeks ahead of the students. It was alot of work for me, but I really enjoyed it.
I also taught a Summer course to train Graduate students and Post Docs. I usually got from 7 - 10 individuals per year. As it was my responsibility to train and supervise anyone in the departments with a demonstrated need for microscopy, it was easiest to teach a group rather than individuals.
Both courses were extremely rewarding. There were always a few individuals who mearly wanted to get by, but there were also those with a serious love for the technique. Cheers,
Doug Keenee
At the University at Albany, State University of New York, we: 1. Teach a combined SEM/TEM class once/year. We also offer an advanced class on image processing. 2. Class size is typically very small (3-5 students). During the late 70s, early 80's the class was typically 5-10 students. With growing interest in structural biology/molecular imaging, we hope enrollment will grow to past levels. 3. No charges beyond tuition. Yet. 4. The class uses equipment at the University-wide EM facility (2 TEMs, 1 SEM), as well as those at the Wadsworth Center, NY State Dept. Health (1 antique SEM, 4 conventional TEMs, and intermediate-voltage TEM, and the 1.2MeV machine). There is plenty of beam time for all.
I'd be interested in your findings. Regards, Sam Bowser -------------------------------- Sam Bowser, Ph.D. Wadsworth Center NY State Department of Health Albany, NY 12201-0509 (518) 473-3856 bowser-at-wadsworth.ph.albany.edu
} 1. How often do you teach a TEM course? I teach it every semester if there is a demand for it by 4 or more students. I would prefer to teach it only once a year
} 2. How many grad/undergrads take it? Enrollment varies. It averages 5 students, mostly graduate students. I have been trying to train undergrad who will stay long enough to work on a research project with a professor before leaving for other endeavors. This has been hindered by the fact that tours were previously arranged by the upper division courses. Lately we have been bringing in the freshman biology class on tours and this has sparked new interest. It is too early to tell if this will produce more students at a lower grade level. Our tours involve working equipment (i.e., slicing microtomes, grids in TEM, samples in SEM, thick stained sections on LM, etc) } 3. Do you charge for consumables or beam time? We receive some class supply money and the service contract is covered by the dept. If we get trained students we can expand our researcher pool by putting students to work with professors. Researchers do pay beam time and consumables, but usually are loath to take the time to train or seek out students for EM.
} 4. Does it make use of a departmental or campus wide facility? If so, } does it interfere with researchers using the facility? Most users have been trained here in the Facility and grumble first as students about researchers time, then as researchers about student time. Generally not a problem, although this is a reflection in part of the limited number of users in the Facility. At lab report time, scope time is a premium, otherwise it is usually available on a walk in basis.
Dr. Steve Barlow EM Facility/Biology Dept. San Diego State University San Diego CA 92182-4614 phone: (619) 594-4523 fax: (619) 594-5676 email: sbarlow-at-sunstroke.sdsu.edu
Dear Tom; In response to your inquire. We (One other professor and myself) teach a course called Anatomy 660: Introduction and practice of Electron microscopy. We teach it every other summer school. It runs for 8 weeks and meets every afternoon from 1:00 to about 4:00. There is a hour to two hour lecture and then a demonstration of what subject the lecture was about. It was designed this way for the student to then start a project and work on it for the remaining 4 weeks, for a 12 week summer session total, using any of the techniques they learned about. They supply their own supplies then during that time. They can use the facility free of charge for that time. We call in about 6 other "experts" on our campus to teach special subjects. ie: Immuno, confocal, steriology, etc. We usually have from 8 to 20 students mostly graduate. one or two advanced undergrads. We use the EM Facility and the hope is to eventually attract new users for the facility. We need more customers. Please let me know what your survey shows. If you have more questions please contact me again. Thanks Grayson Grayson Scott 1300 Univ. Ave Univ. of Wisconsin Electron Microscope Facility Madison, Wisconsin 53706 608-262-2993 Fax 608-262-7306 '''
} 1. How often do you teach a TEM course? We teach one TEM course in the fall and SEM in spring and summer semesters. TEM is not as popular as SEM and is certainly more expensive to run. Its days may be numbered here. SEM is thriving, however.
} 2. How many grad/undergrads take it? We limit the TEM course to 7 and the SEM to 12. Grad students only. There is another course on campus for undergrads but this is lecture and lab/demonstration only. They come to our facility for a 2 hr demo of the EM's. Specimen prep, interpretation, etc is taught by the lecturer in the other course. Really interested students then take our course for more hands on training. This helps to weed out students who are not seriously interested.
} 3. Do you charge for consumables or beam time? $150 to take the course. Covers 10 hr scope use. Students must buy all consumables.
} 4. Does it make use of a departmental or campus wide facility? Campus wide facility
If so, does it interfere with researchers using the facility? We have two levels of scopes,older ones for students and newer ones for researchers. We may stop this in the future as it is too expensive to maintain for the benefits (cash) derived from students.
Contact me if you need more info.
############################################################################ # John J. Bozzola, Director Center for Electron Microscopy 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 ############################################################################ #
Background: "Comprehensive University" (as distinct from "Research University", although I find the appelation redundant); 5200 students 4500 of them undergrads. Master's program only, no PhD offered. Roughly 275 biology majors, 12 biology faculty. Research expected of faculty (roughly 1 paper/2 years), involvement of undergrads in faculty research expected. } } 1. How often do you teach a TEM course? Every other year, although I will train students in SEM at any point that their research project may demand it. 222. How many grad/undergrads take it? Last offering, 14 signed up; nine stuck with it. Two of the nine were grad students. } } 3. Do you charge for consumables or beam time? No. Students taking the course pay a $50 lab fee (lab fees for other courses in the dept. are $15-$25). Students must buy their own fine-point foreceps, lab manual, and photographic paper in excess of 50 sheets. We have no method for charging for beam time; external users are welcome to use the facility if they pay for plate film or polaroids (35mm is free). } } 4. Does it make use of a departmental or campus wide facility? If so, does it interfere with researchers using the facility? The facility is departmental. Given the focus on research that involves undergrads, the course not only doesn't interfere with faculty research, it enhances it. Service contracts are paid out of the departmental base budget, which was increased by the necessary amounts as the microscopes were added.
All-in-all, a pretty cool situation here. HTH Julian Smith III Biology Winthrop University
University of Illinois -at- Urbana-Champaign
I am pleased to respond to your request for information about teaching EM. It has been a dismal situation here for the past 3 years since the campus biotechnology center has taken over the operations of the Center for Electron Microscopy. Biotech refused to support instruction as a part of their policy (that is now under review and may be reversed). What happened was that I made an effort to continue some line of instruction, but I do it without TA's, as an overload assignment without extra compensation, and by taking money from my research funds to pay the CEM/Biotech. for any instructional use of equipment and facilities. The single course now offered is termed: Integrated Electron Microscopy, but has a good deal of other microscopies also included along with microanalysis. Because it was just too much for me, there is no lab, but I have fashioned the course to have weekly demonstration periods. The demos are mostly at the Center for Electron Microscopy, but include sites at other places on campus as well. All lab courses have been dropped. The only way students get instruction is to either do an independent research project with a participating faculty member (not many of them), or to pay the CEM for a staff member to give tutorial assistance. For a student with no background this may be about $1,000 to $2,000. Not many takers.
} 1. How often do you teach a TEM course?
The Integrated EM course is offered once per year--but due to my schedule may need to go on alternate year basis. (Previously, the courses were offered both semesters--though not summers--each year.)
} 2. How many grad/undergrads take it?
We have been enrolling about 20-30 students, mostly grads each time, but this semester we have only 10. One student is an undergrad.
} 3. Do you charge for consumables or beam time?
I do not request a fee from the students since we have demos only. As I indicated above, I pay for the demo time.
} 4. Does it make use of a departmental or campus wide facility?
Yes--although the facility has become more of a service lab than a multi-user facility.
If so, } does it interfere with researchers using the facility?
Not to my knowledge. There is plenty of available time on the equipment.
} If you e-mail me directly I will be happy to post a summary of the } responses to the list once they are all in.
} Thanks for your input.
You are welcome.
************************** Richard F. E. Crang Professor of Plant Biology University of Illinois (217) 244-3143 **************************
} 1. How often do you teach a TEM course? Once per Fall.
} 2. How many grad/undergrads take it? About one dozen.
} 3. Do you charge for consumables or beam time? No, but typically users initiate long term microscopial investigations that do generate "U revenue."
} 4. Does it make use of a departmental or campus wide facility? If so, } does it interfere with researchers using the facility? Departmental. Two lab sessions meet on Thursdays. I suspect a poll of current users would reveal that this constitutes minimal interference. We work every day, all hours anyway. Of course lab time is coordinated with two one hour lectures per week, and a dedicated microscopy education computer network.
Best regards,
Mark T. Biedrzycki Computer Networking Laboratory Materials Science and for Microscopy Education (CNLME) Engineering Department at the University of Arizona, Tucson
1. How often do you teach a TEM course? ==} Basic SEM and TEM undergraduate course taught in fall semester each year. Advanced TEM taught every other year in the spring semester, alternating with Advanced SEM and X-ray Microanalysis.
2. How many grad/undergrads take it? ==} 15-30 students take the undergrad SEM/TEM course, aand perhaps half of these are actually graduate students. About 10 graduate students take the Advanced TEM course and the Advanced SEM and X-ray Microanalysis course.
3. Do you charge for consumables or beam time? ==} Both are charged to the courses making them rather expensive to run since each course has approximately 10 different laboratories.
4. Does it make use of a departmental or campus-wide facility? If so, does it interfere with researchers using the facility? ==} All electron microscopes are in the campus-wide central facility located in the Dept. of Materials Science and Engineering. All departments including biology and geology use the instruments in this department. In addition to our six research electron optics instruments, we maintain three 1970s-vintage microscopes for use by our undergraduate and graduate students: a Philips EM301 TEM and two ETEC Autoscan SEMs.
CE Lyman
Charles E. Lyman Phone: (610) 758-4249 Prof. of Materials Science and Engineering Fax: (610) 758-4244 Lehigh University E-mail: cel1-at-lehigh.edu 5 East Packer Avenue Bethlehem, PA 18015-3195
Thomas E. Phillips, Ph.D. Associate Professor of Biological Sciences Director, Molecular Cytology Core Facility 3 Tucker Hall University of Missouri Columbia, MO 65211 (314)-882-4712 (voice) (314)-882-0123 (fax)
Probably the most comprehensive general introduction to immunogold labeling, since it covers most applications, is vol. 1 and 2 from the series "Colloidal Gold: Principles, Methods and Applications" edited by M. A. Hayat (Academic Press, San Diego, CA; 1989 (Vol. 1 and 2) and 1991 (Vol. 3).
Probably the most comprehensive general introduction to immunogold labeling, since it covers most applications, is vol. 1 and 2 from the series "Colloidal Gold: Principles, Methods and Applications" edited by M. A. Hayat (Academic Press, San Diego, CA; 1989 (Vol. 1 and 2) and 1991 (Vol. 3).
Senior technician in biological light (confocal) and electron microscopy including thin sectioning, various replica techniques, and maintainance of extensive array of preparative equipment. Photography and image processing are not part of job. Annual travel to satellite laboratories necessary. Inquire privately to: treese-at-mbl.edu (Tom Reese)
On the continuing discussion of "strange" BSE contrast, especially at low KV:
There is a slight energy dependence (i.e. accelerating voltage) of the backscattered-electron coefficient (eta) as a function of atomic number at 15-30 KV. For example, see figures 3.13 through 3.15 in the Goldstein text (Scanning Electron Microscopy and X-ray Microanalysis, but hey everybody has this right?).
As you know, the atomic number dependence of eta is strongest over the low atomic number range, so that is where BSE contrast is greatest. It has also been pointed out (MAS talks in New Orleans or Breckenridge) that eta may show strong KV dependence as well, especially at lower KV. Compare the values of eta for 5 KV with those for the other values; they are the highest for low atomic number and the lowest at higher atomic number (figure 3.15). There also appear to be some values of eta that exhibit contrast reversal (see fig 3.14). So you could in fact see contrast reversal when comparing BSE intensities between different accelerating voltages.
As others have suggested, the low KV regime is affected by contamination, and since we are really talking about surface analysis when at 5 KV, it is certainly possible to see contrast reversal due to the competing effects of surface properties coupled with the (as yet imperfectly understood) effects of eta dependence on KV.
I'm sorry I don't have the original post, but I recall we are talking about alloys in the B-Si-Mo system. Despite the best of sample prep methods, it may be that there is surface oxidation (or some other reaction product) that is making itself known to you at 5 KV that was not as important at 15-20 KV.
It certainly is an interesting aspect of microanalysis.
Paul Carpenter
+----------------------------------------------------------------------+ | Paul K. Carpenter paulc-at-arms.gps.caltech.edu | | Division Analytical Facility Department of Geology 170-25 | | California Institute of Technology Pasadena CA 91125 | | 818-395-6126 (X-ray Lab) 818-568-0935 (FAX, Departmental) | +----------------------------------------------------------------------+
This is a summary of the responses I received to my original request (included below) for information on polymer nanoparticle sample prep. =20
Thanks to all who responded!
Owen
} =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D } } Hello; } } I'm working with a group that wishes to make morphological measurements on polymer nanoparticles using electron microscopy (SEM or TEM). The nanoparticles will likely be polyvinylpyridine, polycyanoacrylate, possibly polymethylmethacrylate. The nanoparticles should be in the range of 100-200nm, but they may well be bigger. They would like to see the whole range of particles and be able to report average particle diameter and the whole range of particle sizes (size polydispersity). =20 } } Previous attempts, by placing drops of particles suspended a solvent on a SEM mount or TEM grid, have resulted in "clumps" of particles that are difficult to measure. =20 } } Can anyone recommend a prodecure that might work for this situation. I have heard of (but am unfamiliar with) aerosol procedures, will that work? } } Also, can anyone recommend a few useful general references on electron microscopy of polymers. I have "Polymer Microscopy" by L. Sawyer & D. Grubb on order now. } } Thanks. } } Owen Mills } =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D } } The nanoparticles should be in the range of } } 100-200nm, but they may well be bigger. They would like to see the whole } } range of particles and be able to report average particle diameter and the } } whole range of particle sizes (size polydispersity). =20 } } I have observed some nanoparticles in TEM to obtain the highest contrast= for } automatic image analysis. In SEM the measurements are not as easy as in= TEM. } } } } } Previous attempts, by placing drops of particles suspended a solvent on a } } SEM mount or TEM grid, have resulted in "clumps" of particles that are } } difficult to measure.=20 } } We have observed the same phenomena, too. If your support film can stand= it, } try using e.g. ethanol as the liquid for the suspension. In our case it } helps to keep the particles better in a "monolayer" but we have had } difficulties with the TEM-support films - they trend to break due to= ethanol } (they stand nicely water). } =20 } } } } Can anyone recommend a prodecure that might work for this situation. I= have } } heard of (but am unfamiliar with) aerosol procedures, will that work? } } Here I cannot help you but I know that e.g. virus particles, DNA, etc. can } be spread by aerosol blowing. Try to find the method in e.g. "Practical } methods in electron microscopy". } } } } Let me hear about your success. } } } Best wishes, } } Jouko M=E4ki } Jouko K. M=E4ki, Ph.D., Laboratory Manager } University of Turku, Laboratory of Electron Microscopy } Kiinamyllynkatu 10 FIN-20520 TURKU FINLAND } Tel: +358 21 633 7318 GSM: +358 40 505 2521 FAX: +358 21 633 7380 } } =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D } 1. use some surfactant (Triton-X?) in low concentration in solution with= the } particles to help reduce clumping - the clumping is basically a surface } tension problem and any ting you can do to reduce the surface tension will } help.=20 } } 2. Try serial dilutions of the particles. } } 3. Disperse them onto 12mm glass cover slips for SEM or carbon films for= TEM } - you might try altering the surface characteristics of these substrates = to } help dispersion (glow discharge, chemical treatment, etc.) } } 4. Atomization will help but for such small particles you will probably= just } make micro clumps but it is easy to do so worth trying.=20 } } Good Luck - } } Bill Miller } } =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D A paper, "Quantification of particle sizes with metal replication under= standard } freeze-etching conditions: a gold ball standard for calibrating shadow= widths } was used to measure freeze-etched globular proteins" Microscopy research= and } Technique 32 (1995) 312-329.] was just published that employs 45deg. } unidirectional Pt-C replication to measure particles. This paper will give= you } insight into the problems of replication and how to avoid the pitfalls----= at } the end of the paper is a vertical replication method which (Fig. 17b) is= the } most powerful and reliable of the replication methods. This method employs= a } small metal coating correction to achieve the actual particle size (see ref= in } paper). This method would allow you to measure the asymmetry of the= particles } as well.=20 } Another method which is more readily available to you but can not help } you in the 1-2 nm size range is a negative staining technique. For this= method } you would need to make up a 2% solution of Uranyl acetate in a tin foil= covered } bottle (light sensitive) and filter thru a 0.22 micron filter before use!= You } would need 10-12 nm thick carbon films covered 300 mesh grids. Use a 10 } microliter drop of your particles in ethanol, propyl or isoppropyl alcohol= or } some solvent compatible with water ---- and put it on the carbon film for a } minute. Remove the excess alcohol from the edge of the grid with torn= Whatman } #1 filter paper. Apply 5-10 microliters of stain for a minute and remove= with } the filter paper method and repeat the procedure! Dry the grids from the= edge } with torn Whatman #1 filter paper and then look at your grids in the TEM.= You } will want to measure the white particles you see surrounded by dark stain. = =20 } } George C. Ruben } Dept. Biological Sciences } Dartmouth College } Hanover, NH 03755 } 603-646-2144 } 603-646-1347 fax } George.C.Ruben-at-Dartmouth.EDU } } =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D } We do this type of analysis by cryoTEM of vitrified thin liquid films of } dispersed particles. We collect the images on a slow-scan CCD camera using } Gatan's DigitalMicrograph and do image analysis using NIH Image. } We compute the size distributions using a statistical package } called RS/1 (on the VAX or Sun) and fit the data to a lognormal } distribution with up to three modes. } } We have tried aerosol methods and have not been satisfied with the } fraction of isolated particles. We do MUCH better with cryoTEM. } The good news is that one can automatically } select only isolated spheroidal particles from the images } containing some overlapping particles by calculating } the circularity of each feature (C=3D4*pi*area/perimeter**2). For such= particles } we only keep features with 0.90 } C } 1.05. Verify this yourself by } measuring model images of agglomerated circles. By the way, it took me } almost a year to develop this capability to the point that I convinced } myself that I had done everything correctly. It was not a trivial } project. } } minter-at-Kodak.COM (John Minter) } =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D } I don't know if my suggestion will be of any use but I guess it's worth=20 } sharing. I work with nanometre size ceramic particles and to get them in= =20 } the TEM I disperse them in methanol ultrasonically and then pick them up on= =20 } a carbon film supported on a copper grid. I then place the grid on some=20 } tissue or filter paper which quickly wicks away the methanol. This usually= =20 } gives good results if the particle suspension is thin enough. We usually= =20 } see some particle overlaps but there are usually enough free particles to= do=20 } some good TEM. } Do you use ultrasonic agitation to break up any particle agglomerates (we= =20 } need to as ceramic systems are often very prone to agglomeration)? What=20 } liquid do you use to disperse the particles in? Have you tried all the=20 } ideas suggested above already? } } I hope this is of some help to you } } Best wishes } } } Ian MacLaren, } IRC in Materials for High Telephone: 0121 414 3447 =20 } Performance Applications, FAX: 0121 414 3441 } The University of Birmingham, email: I.MacLaren-at-bham.ac.uk } Birmingham B15 2TT, =20 } England } =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D } In my prior life as a chemical engineer, I was responsible for a colloidal } silica unit. Whenever questions were raised about particle size } distribution, we would walk a sample over to our friendly electron } microscopist for analysis. } } His secret was to perform a series of dilutions to very dilute levels of= Si. } My recollection is that he did a series of three or four 1:100 dilutions } before drying a drop on a coated grid. This created a single particle= thick } film of these normally sticky SiOx particles. } } We were routinely dealing with discrete particle sizes from 6 to 150 nm. } } } Good luck, } Joe Tabeling } Delaware Diamond Knives } 800-222-5143 } } =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D } 1) You may have to try to sonicate the suspension right before=20 } putting it on the grid or SEM stub. } =09 } 2) If you can get hold of the dry sample (before they put it in=20 } the solvent) and if it looks powdery then you can just pick a little bit=20 } up with the tip of a pastuer pipette and gently blow it on the grid or=20 } glass cover slid for SEM. } } } Have fun and good luck. } } Yew Meng Heng } } E.M. Facility=20 } Mcmaster University Medical Centre } Hamilton, Ont., Canada } =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D } -- [ From: Charles A. Garber, Ph. D. * EMC.Ver #2.10P ] -- } } On Nov. 7, Owen P. Mills wrote inquiring about the preparation of } polymer particles in the range of 100-200 nm. } } We do a large volume of these kinds of samples in our laboratory: } } 1] If they are clumping, then probably the concentration of the } suspension is too high and you need a higher dilution by at least one } and maybe two or three orders of magnitude. } } 2] You want to use an "aerosol" type "duster" valve that incorporates } a capillary tube to disperse the now highly diluted suspension. The } best (but certainly not the only) one I know of is made by Ernest F. } Fullam, Inc. in Schenectady, NY. The cost is less than $100. This is } in all probability the "aerosol" method you mentioned in your posting. } } 3] You have to worry about the glass transition temperature. If above } room temperature, then you need not further worry. If below room } temperature, then the particles at room temperature will be soft and } you have to worry about the need to "harden" them up. Acrylic } particles can be "hardened" for example by UV using quartz irradiation } tubes. } } 4] While particles in this range could in theory be done by SEM, } especially cryo SEM, you will find that Pt/C replication TEM will give } much more acceptable results. Use a shadowing angle of 45 degrees.=20 } You can measure the "shadow" and also the apparent diameter, and then } compare the measurements as a insight into any "collapse" of the } particles, being sort of a built in validation that the particles are } indeed hardened into "hard" rigid marbles. } } You do not need anything fancy in the way of a vacuum evaporator, any } system giving a vacuum down in the "mid to low 10 to the minus five" } range will give acceptable results.=20 } } 5] If graft polymer latex, then if you want to see the morphological } details of the graft vs. substrate polymer phase, you will have to use } a still different technique to bring out such structures. But again, } the method actually used will be determined by the specific polymers } present and the kind of reactivity they exhibit. } } Chuck } } =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D } 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: } GVKM07A-at-prodigy.com } West Chester, PA 19381-0656 USA Customer Service:=20 } SpiSupp-at-aol.com } =20 } ######################################################## =20 } WWW: http://mail.cccbi.chester.pa.us/spi/spihome.html =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D As Dr Charles A Garber wrote there are special problems to image polymers in TEM. If you are not a polymer scientist, the negative staining method is the most safe one. It will give a proper result even if you have a film forming (low glass transition) or beam sensitive polymer. When you have learnt to dilute the latexes and add appropriate amount of Uranyl Acetate or PTA, it is also very easy to use.
Helen Hassander Polymer group, Lund University
=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D=3D= =3D=3D=3D=3D=3D=3D=3D There is an article in the latest issue of THe Microscope that might be of= =20 interest: "Cryo-Ultramicrotomy of Individual Latex Particles for Examination= =20 of Internal Morphology", Marcelli, Angela. The Microscope= 43(3):117-120(1995)
Peter D. Barnett Forensic Science Associates e-mail: pbarnett-at-crl.com Owen P. Mills Michigan Technological University Metallurgical & Materials Engineering Rm 512 MME Building Houghton, MI 49931 906-487-2002 906-487-2934 FAX opmills-at-mtu.edu
It might be a good idea to directly look at the BSE signal levels coming from different parts of the sample rather than the differences of those signals. Just to be sure that the 'image contrast reversal' isn't a product of the electronics.
I have been contacted by an R&D group for a Pharmaceutical manufacturing firm. They are interested in the integrity of the seal on gelatin capsules. They haven't decided exactly in which direction to take the project, but there are several possibilities:
1. Frozen sections, 1-5 microns thick, across the seal, for light microscopy/F.T.I.R., etc.
2. Cryoultramicrotomy for EM of the seal
3. cryoultramicrotomy of seal area followed by SEM of the sectioned surface.
My schedule won't permit me to do the work. Is there anyone (preferably in the southeastern US) who would like to work on this?
The R&D group is commited to doing the project, and they have a budget for the work.
If you are interested, please contact me at rmcbtli-at-aol.comb (not the address above) for additional information.