Date: Wed, 1 Nov 1995 12:57:41 +0000 (GMT) From: Ray Hicks Subject: Re: Microwave fixation 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. Ray. ============================================================================== Date: Wed, 1 Nov 1995 23:34:46 -0400 From: jfmjfm@umich.edu (John F. Mansfield) Subject: Re: Microwave fixation, actually continued comment 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. 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@engin.umich.edu, John.F.Mansfield@umich.edu or jfmjfm@umich.edu they all reach me! URL: http://wwwpersonal.engin.umich.edu/~jfmjfm/jfmjfm.html ============================================================================== Date: Wed, 1 Nov 1995 13:19:12 -0700 From: chandler@lamar.ColoState.EDU (John Chandler) Subject: Re: Microwave fixation 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. John chandler@lamar.ColoState.EDU ============================================================================== From: BARBARA.HARTMAN@1773.220.SCHERING-PLOUGH.sprint.com Date: Wed, 1 Nov 1995 12:16:33 -0500 Subject: Uranyl acetate and lead citrate disposal Greetings: 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 E-Mail: Barbara.Hartman@Schering-Plough.sprint.com ============================================================================== Date: 1 Nov 1995 15:11:42 -0400 From: "Paul Webster" Subject: Re: reduced osmium 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 ============================================================================== Date: Thu, 2 Nov 1995 07:59:42 -0500 From: rjpalmer@utkux.utcc.utk.edu (Robert J. Palmer Jr.) Subject: Re: Microwave fixation, actually continued comment 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! ============================================================================== Date: Thu, 2 Nov 1995 08:56:28 -0600 From: baskin@biosci.mbp.missouri.edu (Tobias Baskin) Subject: Re: Microwave fixation, actually continued comment 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 ============================================================================== From: trenkler@imec.be Date: Thu, 2 Nov 1995 21:46:51 +0100 Subject: re:PAL LAserdisc to sth. extract from faq@soc.cult.german I know, it's not about laser disks, but give it a try! Subject: 8 Audio / Video Tapes =============================== Subject: 8.1 Different Video Norms! ------------------------------------ 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@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] ============================================================================== Date: Thu, 2 Nov 1995 14:02:49 GMT From: gwe@biotech.ufl.edu (Greg Erdos) Subject: Re: Microwave fixation, actually continued comment 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 @@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@ 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@biotech.ufl.edu Gainesville, FL 32611 @@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@ ============================================================================== From: "IAN HALLETT" Organization: HortResearch, Mt Albert, N.Z. Date: Fri, 3 Nov 1995 09:13:50 GMT+1200 Subject: Re: Microwave fixation, actually continued comment In reply to John Mansfields remarks 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 Hallett HortResearch Mt Albert Research Centre Private Bag 92 169 Auckland, New Zealand Fax 64-9-815 4201 Telephone 64-9-849 3660 ============================================================================== Date: Sat, 4 Nov 1995 03:24:25 -0600 From: bozzola@siu.edu (John. J. Bozzola) Subject: CSMS-MIKMAS Meeting in St. Louis CSMS-MIKMAS PROGRAM 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@siu.edu Web: http://www.siu.edu/departments/shops/cem.html ############################################################################# ============================================================================== Date: Sat, 4 Nov 1995 14:22:06 -0500 (EST) From: "Carmine M. Pariante" Subject: quantitative immuno-fluorescence Dear collegues, 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@emory.edu ============================================================================== Date: Tue, 7 Nov 1995 13:18:27 +0000 From: deschuyt@ccmailg1.sbbio.be (Michel DESCHUYTENEER) Subject: Postings and replies, a technical note Greetings everyone. 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@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 ============================================================================== Date: Tue, 7 Nov 1995 17:38:09 -0500 From: "Owen P. Mills" Subject: Sample prep - polymer nanoparticles 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@mtu.edu ============================================================================== Date: Tue, 7 Nov 1995 23:01:20 -0600 From: zaluzec@Microscopy.Com (Nestor J. Zaluzec) Subject: : High Resolution EDS Detectors for Microanalysis Don Morgan asked........ 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. Cheers... Nestor Your Friendly Neighborhood SysOp ============================================================================== Date: 08 Nov 95 07:25:27 EST From: George.C.Ruben@Dartmouth.EDU (George C. Ruben) Subject: Re: Sample prep - polymer nanoparticles Dear Owen, 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@Dartmouth.EDU ============================================================================== Date: Wed, 08 Nov 1995 08:45:33 -0600 From: wesaia@iastate.edu (Warren Straszheim) Subject: Re: Sample preparation 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@ameslab.gov (or: wesaia@iastate.edu) coal characterization and processing electron microscopy, x-ray analysis, image analysis computer applications ============================================================================== Date: Wed, 8 Nov 1995 09:56:54 -0500 From: minter@kodak.com (John Minter) Subject: Re: Sample prep - polymer nanoparticles 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. ============================================================================== Date: Wed, 8 Nov 1995 09:52:58 -0700 From: Jeffrey.Shield@mse.utah.edu (Jeff Shield) Subject: JEOL2000FX 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 ============================================================================== Date: Wed, 8 Nov 1995 12:52:14 -0500 From: Peters@BSAC.UCHC.EDU (Klaus-Ruediger Peters) Subject: Re: Microscopical demo-images site 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. Thanks Klaus ****************************************************************************** * : * * Klaus-Ruediger Peters, Ph.D. : Browse our WWW Home Pages: * * Director, Molecular Imaging Laboratgory : * * Biomolecular Structure Analysis Center : Molecular Imaging Laboratory * * University of Connecticut Health Center : http://panda.uchc.edu/ * * 263 Farmington Ave. : htklaus/index.html * * Farmington, CT 06030-2017; U.S.A : Differential Hysteresis * * : Processing Demo at http:// * * Tel: (203) 679-3977; Fax: (203) 679-1989 : panda.uchc.edu./htbit/indiv/ * * e-mail: Peters@BSAC.UCHC.EDU : /software_docs/dhp.html * * : * **************************************************************************** ============================================================================== Date: Wed, 8 Nov 1995 10:43:26 -0600 From: "Gib Ahlstrand" Subject: Re: SEM Protocol for Insect Eyes In message 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@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@puccini.crl.umn.edu "MICROSCOPY & MICROANALYSIS 96" in Minneapolis, Minnesota, Aug.11-15, 1996 ============================================================================== Date: Wed, 8 Nov 1995 16:52:05 -0800 (PST) From: Michael Rock Subject: Re: digital SEMs 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@conncoll.edu ============================================================================== Date: Wed, 8 Nov 1995 16:11:54 EST From: pat_masarachia@Merck.Com (Pat Masarachia) 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 ============================================================================== From: William Tivol Subject: Re: High Resolution Microanalysis Date: Wed, 8 Nov 1995 17:39:02 -0500 (EST) 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 ============================================================================== Date: Wed, 08 Nov 1995 11:03:34 -0600 From: "James C. Long" Subject: Voluteers Needed - Science by Mail Program 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@bga.com ============================================================================== Date: Wed, 8 Nov 1995 20:55:43 -0800 From: mager@unixg.ubc.ca (Mary Mager) Subject: Re: Digital SEMs 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@unixg.ubc.ca ============================================================================== Date: Thu, 9 Nov 1995 15:58:10 +1100 From: m.blackford@ansto.gov.au (Mark Blackford) Subject: Re: JEOL2000FX 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 ============================================================================== Date: Thu, 09 Nov 1995 00:04:54 EST From: GVKM07A@PRODIGY.COM (DR CHARLES A GARBER) Subject: TEM Sample Preparation -- [ 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@prodigy.com West Chester, PA 19381-0656 USA Customer Service: SpiSupp@aol.com ######################################################## WWW: http://mail.cccbi.chester.pa.us/spi/spihome.html ######################################################## ====================================================== ============================================================================== Date: Thu, 9 Nov 1995 16:23:26 +1100 From: David Dryden Subject: Re: JEOL 2000FX 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 @ 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@electron.ph.unimelb.edu.au http://www.ph.unimelb.edu.au/~djd/diff-home.html ============================================================================== Date: Thu, 9 Nov 1995 17:44:43 NZS From: Stephen Edgar Subject: Re: Keeping absolute ethanol dry - a summary Hi everyone, 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 email address: s.edgar@auckland.ac.nz Phone : +64-9-3737599 extn 6473 (GMT + 12h) Fax : +64-9-3737459 ============================================================================== Date: 09 Nov 95 07:53:43 EST From: George.C.Ruben@Dartmouth.EDU (George C. Ruben) Subject: Re: slot grid problems 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???????????????????????? Thanks, ============================================================================== Date: Thu, 9 Nov 1995 08:39:08 GMT From: gwe@biotech.ufl.edu (Greg Erdos) Subject: Re: slot grid problems 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 Phone: 904-392-1295 Scientific Director, ICBR Electron Microscopy Core Lab 218 Carr Hall Fax 904-846-0251 University of Florida E-mail: gwe@biotech.ufl.edu Gainesville, FL 32611 @@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@ ============================================================================== From: "Richard E. Edelmann" Organization: Miami University Date: Thu, 9 Nov 1995 13:19:45 -500 Subject: Re: Digital SEMs 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 ? ============================================================================== Date: 9 Nov 1995 16:06:35 -0400 From: "Paul Webster" Subject: Re: LR White Immunolabeling 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 ============================================================================== From: William Tivol Subject: Re: slot grid problems Date: Thu, 9 Nov 1995 17:49:33 -0500 (EST) Dear Shaun, ... 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 Message-ID: Date: 9 Nov 1995 13:26:08 -0600 From: "Richard Lee" Subject: Re: SEM education To: "Michael Odum" , "Microscopy@Sparc5.Microscopy.Co" Cc: "Nancy Daerr" X-Mailer: Mail*Link SMTP-QM 3.0.2 RE>>SEM education 11/9/95 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. ============================================================================== Date: Thu, 9 Nov 1995 09:14:32 -0700 From: modum@gatan.com (Michael Odum) Subject: Re: SEM education 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 @gatan.com ============================================================================== Date: Thu, 9 Nov 1995 22:05:33 -0600 From: jbpawley@facstaff.wisc.edu (James Pawley) Subject: Re: Digital SEMs 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 Prof. James B. Pawley, Ph. 608-263-3147 Room 1235, Engineering Research Building, FAX 608-265-5315 1500 Johnson Dr., Madison, WI, 53706 JBPAWLEY@FACSTAFF.WISC.EDU ============================================================================== Date: Fri, 10 Nov 1995 03:51:22 EST From: GVKM07A@PRODIGY.COM (DR CHARLES A GARBER) Subject: Sample Prep - Polymer Nanoparticles -- [ 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@prodigy.com West Chester, PA 19381-0656 USA Customer Service: SpiSupp@aol.com ######################################################## WWW: http://mail.cccbi.chester.pa.us/spi/spihome.html ######################################################## ====================================================== ============================================================================== Date: Fri, 10 Nov 1995 08:45:41 -0600 (CST) From: Richard Gordon Subject: University of Manitoba Professors Academic Freedom Strike Settled 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 . 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@xpressnet.com, to President Arnold_Naimark@UManitoba.ca, and to Premier Gary Filmon: premier@leg.gov.mb.ca UMFA = University of Manitoba Faculty Association ============================================================================== Date: 10 Nov 1995 09:33:48 -0800 From: "Doug Davis" Subject: Re: Slot grid substrates ... 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. Good luck! doug_davis@maillink.berkeley.edu ============================================================================== Date: Mon, 13 Nov 1995 01:37:49 EST From: GVKM07A@PRODIGY.COM (DR CHARLES A GARBER) Subject: CPD Tissue Baskets -- [ 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@prodigy.com West Chester, PA 19381-0656 USA Customer Service: SpiSupp@aol.com ######################################################## WWW: http://mail.cccbi.chester.pa.us/spi/spihome.html ######################################################## ====================================================== ============================================================================== From: STEELE@KRDC.INT.ALCAN.CA Date: Mon, 13 Nov 1995 14:17:43 -0500 (EST) Subject: EPOL TEM X-sections --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@KRDC.INT.ALCAN.CA # # ALCAN INTERNATIONAL # # Kingston Research and Development Center # # P. O. Box 8400 # # Kingston, Ontario Canada K7L 5L9 # # # ############################################################### ============================================================================== From: "Rolf Odselius" Organization: Lund University Date: Mon, 13 Nov 1995 10:11:26 +0100 Subject: Change of WWW address! The address to the WWW pages of the Electron Microscopy Unit Medical Faculty Lund University SWEDEN has changed (again!). I beleive that this new address will keep for several years. The old URL was: http://www.medfak.lu.se/medinst/emu/ ____________________________ | | | THE NEW URL IS: | | | | http://www.emu.lu.se/ | |____________________________| Please change your address books and links. Sorry for the inconveniance! With best regards Rolf Odselius ____________________________________________________________________ Rolf Odselius, PhD |Rolf.Odselius@emu.lu.se Electron Microscopy Unit |Phone: +46 46 171075 office University Hospital | +46 46 171155 lab S-221 85 Lund, Sweden | +46 46 293692 home | +46 10 6705655 mobile http://www.medfak.lu.se/medinst/emu/ |Pager: +46 740 288992 SCANDEM: http://www.ldc.lu.se/~scandem/|Fax: +46 46 172975 ============================================================================== Date: Sun, 12 Nov 1995 17:51:26 -0600 From: Colin Veitch Subject: EELS Simulation Hi all, 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@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." # # # ############################################################################### ============================================================================== Date: Sun, 12 Nov 1995 18:20:13 -0300 (BST) From: Adriana Pinheiro Martinelli Rodriguez Subject: Re: mounting media 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 ============================================================================== Date: Sat, 11 Nov 1995 18:57:45 -0800 (PST) From: Daniel Possin Subject: Re: LR White immunolabeling-LM/EM Hi Pat - 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 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@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". % % % %%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%% ============================================================================== Date: Mon, 13 Nov 1995 18:02:13 -0500 From: Benjamin Walcott Subject: Re: LM/ Hg HBO 50W lifetimes In , 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 ============================================================================== Date: Sat, 11 Nov 1995 11:32:53 EST From: GVKM07A@PRODIGY.COM (DR CHARLES A GARBER) Subject: JEOL 2000FX arcing -- [ 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@prodigy.com West Chester, PA 19381-0656 USA Customer Service: SpiSupp@aol.com ######################################################## WWW: http://mail.cccbi.chester.pa.us/spi/spihome.html ######################################################## ====================================================== ============================================================================== Date: Mon, 13 Nov 1995 13:12:39 -0500 (EST) From: Nina S Allen Subject: Re: confocal vs deconvolution microscopy 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? Nina Allen ============================================================================== Date: Mon, 13 Nov 1995 15:21:35 -0600 (CST) From: jr Subject: Re: SEM education 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 @gatan.com ============================================================================== From: William Tivol Subject: Re: LM/ Hg HBO 50W lifetimes Date: Mon, 13 Nov 1995 15:03:20 -0500 (EST) 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 ============================================================================== Date: Sun, 12 Nov 95 17:52:48 0100 From: "Klaus F. Oestergaard" Subject: LM spares for Olympus VANOX - light source Hello Microscopists, We have an old Olympus VANOX microscope, fitted only with reflected light. Now wee wants to do both reflected and transmitted light microscopy, the microscope is prepared for it, but we are missing a light source and a condenser system before we are able to do transmitted light microscopy. We have not been able to find any local dealers their can help us, maybe their are some people on this listserver who can help us - both contacts, new parts and used parts have interest. Thanks, Klaus, PS. Please send mail direct to me. -------------------------------------------------------------------------- Klaus F. Oestergaard Department of Physics Telephone +45 4525 3111 Technical University of Denmark Direct +45 4525 3118 Building 307 Fax +45 4593 2766 DK-2800 Lyngby Denmark e-mail klaus@carbon60.fysik.dtu.dk URL: http://carbon60.fysik.dtu.dk:8080/~klaus --ooo00ooo-- -------------------------------------------------------------------------- ============================================================================== Date: Sat, 11 Nov 1995 19:15:49 -0800 (PST) From: Daniel Possin Subject: Re: slot grid problems Hi Shaun - 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 To: microscopy@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@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". % % % %%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%% ============================================================================== Date: Fri, 10 Nov 1995 19:24:57 -0600 From: jbpawley@facstaff.wisc.edu (Jim Pawley) Subject: Re: Digital SEMs Dear Warren, 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@FACSTAFF.WISC.EDU ============================================================================== Date: Tue, 14 Nov 1995 08:17:45 -0600 From: "Richard W. Fonda" Subject: Re: Edington TEM Monographs 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@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@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 _____________________________________________________________ ============================================================================== Date: Tue, 14 Nov 1995 09:03:01 -0500 (EST) From: colijn@kcgl1.eng.ohio-state.edu (Henk Colijn) Subject: Re: Edington TEM Monographs You can get the book from Tech Books, Inc, 2600 Seskey Glen Ct. Herndon, VA 22071 (703) 758-1518 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@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@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. ============================================================================== From: !Microscopy-request@Sparc5.Microscopy.Com (Robert Kayton,MAC,CROET) Date: Sat Oct 28 21:18:41 GMT 1995 Subject: Microscopy Manual Online 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 - America - 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@stowey.demon.co.uk | | (work) - chris.jefferies@bbsrc.ac.uk | | Microscopy page (UK) | | (USA) | ----------------------------------------------------------------------------- ============================================================================== From: !Microscopy-request@Sparc5.Microscopy.Com (Albert Romano Rodriguez) Date: Fri Oct 27 08:49:26 +0100 95 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@iris1.fae.ub.es tel: +34 3 402 11 47 FAX: +34 3 402 11 48 ============================================================================== From: !Microscopy-request@Sparc5.Microscopy.Com (Robert Kayton,MAC,CROET) Date: Fri Oct 27 15:52:22 -0400 1995 Subject: More on Lexmark Optra R 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 ============================================================================== From: !Microscopy-request@Sparc5.Microscopy.Com (Robert Kayton,MAC,CROET) Date: Mon Oct 30 16:46:44 -0500 1995 Subject: Re: Microwave fixation 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 glogin@ bih.harvard.edu Telephone: 617-667-2034 Fax: 617-667-8676 ============================================================================== Date: Wed, 15 Nov 1995 08:38:39 -0800 (PST) From: Michael Shaffer Subject: Re: 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@oregon.uoregon.edu -or- mshaf@darkwing.uoregon.edu http://darkwing.uoregon.edu/~mshaf/epmahome/ ============================================================================== Date: Wed, 15 Nov 1995 12:21:39 -0500 (EST) From: DALE A CALLAHAM Subject: Re: Arc Lamps **************************************************************** 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 (@ 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 email: dac@bio.umass.edu ============================================================================== From: "Greg" Date: Wed, 15 Nov 1995 13:05:00 EST5DST Subject: Re: 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@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@cle.dnet.ge.com ============================================================================== Date: 15 Nov 1995 10:57:28 -0800 From: "Murphy, Judy" Subject: RE: Microscopy Manual Online 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@stowey.demon.co.uk on Wed, Nov 15, 1995 10:22 AM Subject: Microscopy Manual Online 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 - America - 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@stowey.demon.co.uk | | (work) - chris.jefferies@bbsrc.ac.uk | | Microscopy page (UK) | | (USA) | ----------------------------------------------------------------------------- ============================================================================== Date: Wed, 15 Nov 1995 10:21:02 -0800 (PST) From: sbarlow@sunstroke.sdsu.edu (Steve Barlow) Subject: EM:Workshop on microwave fixation 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@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 demonstr