I'm doing a bit of testing right now. Hopefully you will not notice any problems, however, if you do please send an error report directly to:
Zaluzec-at-ANLEMC.MSD.ANL.GOV
I'm in the process of slowly upgrading the listserver software abit at a time to get rid of some of the annoying problems that have been creeping into the system. and the subsequent headaches for you and me. This process will likely take awhile, and I will be switching periodically between old and new links. Most of the installation was done over the holidays however, I've still got a load of reconfigurations which need to be handled "gently" to minimize any crashes. However, as Murphy has it I expect some problems. I'll keep you all informed as I make any major changes, but expect some hickups.
Subject: Time:2:17 PM OFFICE MEMO Resolved-Focusing on JEOL 1010 Date:1/3/94 Just a short note to thank everyone who responded to my request for help in rectifying our focusing problem with our the JEOL 1010. As I indicated in an earlier message, we learned a lot from the many comments and suggestions by the members of this list. It ends up that the gun alignment was changing when the shutter closed, which caused beam sensitive specimens like we use, i.e. slotted grids, to charge. Evidently, this is not a serious problem with mesh grids as are used in calibration. JEOL indicated that an early design innovation of the 1010 which was intended to blank the beam prior to exposure was later found to be less than satisfactory and abandoned this method in favor of the traditional system. Unfortunately, in our scope some remnants of this circuitry were not eliminated, thus contributing to our problem. They were able to track this problem down after considerable, and diligent effort, and corrected it by modifying the deflector lens amperage and by removing circuit elements linking the shutter and gun alignment. I am not sure what all this means, however, all or photographs since this correction have been in perfect focus. JEOL has been very cooperative and attentive to our frustrations. We are thankful that they did not take the approach of one of the members of this list who indicated that: "This is really simple! If anyone can take sharp pictures using standard specimens then the machine is working fine!" Hopefully, this information will be of use to other users of the JEOL 1010 which may encounter similar focusing problems with beam sensitive specimens.
Subject: Time:2:17 PM OFFICE MEMO Resolved-Focusing on JEOL 1010 Date:1/3/94 Just a short note to thank everyone who responded to my request for help in rectifying our focusing problem with our the JEOL 1010. As I indicated in an earlier message, we learned a lot from the many comments and suggestions by the members of this list. It ends up that the gun alignment was changing when the shutter closed, which caused beam sensitive specimens like we use, i.e. slotted grids, to charge. Evidently, this is not a serious problem with mesh grids as are used in calibration. JEOL indicated that an early design innovation of the 1010 which was intended to blank the beam prior to exposure was later found to be less than satisfactory and abandoned this method in favor of the traditional system. Unfortunately, in our scope some remnants of this circuitry were not eliminated, thus contributing to our problem. They were able to track this problem down after considerable, and diligent effort, and corrected it by modifying the deflector lens amperage and by removing circuit elements linking the shutter and gun alignment. I am not sure what all this means, however, all or photographs since this correction have been in perfect focus. JEOL has been very cooperative and attentive to our frustrations. We are thankful that they did not take the approach of one of the members of this list who indicated that: "This is really simple! If anyone can take sharp pictures using standard specimens then the machine is working fine!" Hopefully, this information will be of use to other users of the JEOL 1010 which may encounter similar focusing problems with beam sensitive specimens.
This newsgroup passed its vote by a significant margin. Barring SERIOUS controversy, it will be created in five days.
Newsgroups line:
sci.techniques.microscopy The field of microscopy.
Voting closed at 23:59:59 UTC, 2 January 1994.
After this result appears on news.announce.newgroups it will be sent to the following mailing lists (minus the individual ACK lists), with the permission of their respective maintainers:
microscopy-at-anlemc.msd.anl.gov {Microscopy Mailing List} maintained by Nestor J. Zaluzec {zaluzec-at-anlemc.msd.anl.gov}
nih-image-at-soils.umn.edu {NIH Image Mailing List} maintained by John Ladwig {jladwig-at-soils.umn.edu}
This vote was being conducted by a neutral third party. For voting questions only, contact pschleck-at-unomaha.edu. For questions about the new group, contact John F. Mansfield {John.F.Mansfield-at-umich.edu} .
(Vote-takers Note: I apologize for the lack of a 2nd CFV. It was my intention to send one out halfway through the voting period, on December 19th. However, I wasn't paying close attention to the news.announce.newgroups submission deadline before the holidays, which came up earlier this year, on December 18th. The extremely decisive results, combined with the usual lull in activity over the holidays, makes it almost impossible that this accidental oversight affected the the outcome of the vote, anyway.)
CHARTER
The main aim of sci.techniques.microscopy is to provide an open forum for the discussion of microscopy and related fields on the Internet.
PURPOSE
The purpose of sci.techniques.microscopy is to provide an open discussion forum for the microscopy community on the Internet. The newsgroups allow the rapid and timely discussion of opinions and information that would take months or years (or not at all) on conventional paper journals. It is hoped that this newsgroup will eventually be linked with the microscopy-at-anlemc.msd.anl.gov mailing list and archived at the same site. Technical suggestions as to the best way to accomplish this are welcome, and may be directed to either John F. Mansfield or Nestor J. Zaluzec via their respective E-mail addresses above.
Please note that this proposed newsgroup is intended to be an open forum for discussion of microscopy. Thus relevant topics for this newsgroup should only be limited to what the participants in this proposed newsgroup regard as microscopy.
TOPICS FOR DISCUSSION
Optical Microscopy Confocal Microscopy Scanning Probe/Force Microscopy (formerly Atomic Force Microscopy) - SPM, SFM, AFM Scanning Tunnelling Microscopy - STM Scanning Electron Microscopy - SEM Transmission Electron Microscopy - TEM High Resolution Electron Microscopy - HREM Analytical Electron Microscopy - AEM Scanning Transmission Electron Microscopy - STEM High Voltage Electron Microscopy - HVEM X-ray Energy Dispersive Spectroscopy - XEDS Electron Energy Loss Spectroscopy - EELS Electron Microprobe Analysis (EMPA) Wavelength Dispersive X-ray Spectroscopy (WDS) Diffraction Contrast Imaging Phase Contrast Imaging Selected Area Electron Diffraction - SAED or SAD Convergent Beam Electron Diffraction - CBED Image Filtering Field Ion Microscopy Electron Holography X-ray Microscopy Scanning Acoustic Microscopy Ultrasonic Imaging Specimen Preparation (Electropolishing, Ion Milling, Ultramicrotoming, etc.) 3D reconstruction Image Processing Software Data formats Databases Hardware/Equipment - specs, opinions, etc. Applications Announcements/reviews of papers/conferences. Preparation techniques. Non-ambient techniques General Discussion/opinions/questions. Positions vacant
As well as anything else that is relevant to microscopy in general.
[sci.techniques.microscopy group vote Final Vote Ack deleted to avoid overloading the mailing lists. The full results may be obtained via anonymous FTP from ftp.uu.net under /usenet/news.announce.newgroups/ sci/sci.techniques.microscopy.]
Hello, I just started working with magnetic steel specimens in the TEM and find that the beam is just "bent" everywhere. Has anyone worked with steel before and/or does anyone have any pointers they picked up along the way? Thanks..
Return-Path: {P.V.Hatton-at-sheffield.ac.uk} Received: from oxmail.ox.ac.uk by vax.ox.ac.uk (MX V3.3 VAX) with SMTP; Tue, 04 Jan 1994 14:43:36 +0000 Received: from sheffield.ac.uk by oxmail.ox.ac.uk via JANET with NIFTP (PP) id {17715-0-at-oxmail.ox.ac.uk} ; Tue, 4 Jan 1994 14:43:26 +0000 Received: from pp2.shef.ac.uk by pp.shef.ac.uk with SMTP (PP) id {04547-4-at-pp.shef.ac.uk} ; Tue, 4 Jan 1994 14:41:54 +0000 Received: from gorgon.shef.ac.uk by pp2.shef.ac.uk with SMTP (PP); Tue, 4 Jan 1994 14:33:35 +0000 Received: From LADYBOWER/GORGON_WORKQ by gorgon.shef.ac.uk via Charon-4.0A-VROOM with IPX id 100.940104143216.2368; 04 Jan 94 14:33:35 +0000 Message-ID: {MAILQUEUE-101.940104142755.448-at-UNDERBANK.shef.ac.uk} To: rms-at-vax.ox.ac.uk
I would be grateful if you could circulate the following info:
"X-Ray Microanalysis - Advances and Applications" The Spring Meeting of the X-ray Microanalysis Group will be held at the School of Clinical Dentistry, University of Sheffield on Thursday 7th April 1994. Enquiries can be directed to Dr Paul Hatton, tel. 0742 670444 ext. 3051, fax 0742 797050
Journal of Microscopy, Vol. 173, Pt. 1, January 1994, pp. 1--8.
The observation of large magnetite crystals from magnetotactic bacteria by electron and atomic force microscopy
by MARCOS FARINA,* BECHARA KACHAR,"" ULYSSES LINS,* RAYMOND BRODERICK~~ & HENRIQUE LINS DE BARROS##,
*Instituto de Biofisica Carlos Chagas Filho - CCS - Bloco G -Universidade Federal do Rio de Janeiro, 21949-900, Rio de Janeiro, Brazil. ""Laboratory of Cellular Biology, National Institute on Deafness and Other Communication Disorders, Bethesda, Maryland, U.S.A. ~~Department of Pharmacology and Experimental Therapeutics, University of Maryland at Baltimore, School of Medicine, Maryland, U.S.A. ##Museu de Astronomia e Ciencias Afins, Rio de Janeiro, Brazil
Summary Magnetite crystals inside coccoid magnetotactic bacteria found in lagoons near Rio de Janeiro city were examined by electron microscopy (EM) and atomic force microscopy (AFM). For AFM, ultrathin sections of bacteria embedded in Epon resin were etched with an ethanolic NaOH solution and observed both in the height and in the force modes. Comparative electron microscope images were useful for identifying crystalline reliefs in the etched sections. Different situations representing particular arrangements of crystal chains were observed by AFM. The majority of the bacteria examined presented unusually large magnetite crystals which remained strongly attached in linear chains even after the laboratory procedures for their isolation. This behaviour is different from all other biogenic magnetite crystals isolated so far. It is suggested that this attachment is due to the strong field between individual crystals as well as to the contact areas, which are the largest observed until now. The correct identification of a particular topography by AFM as a crystal relief may be critical when crystals are not aligned in chains; in these cases the linear dimensions and the presence of well-defined edges and faces are important features to be taken into account. Characterization of the crystal faces is important for the study of magnetotactic micro-organisms since the crystalline habits seem to be species-specific. Observation of etched sections proved to be a helpful approach for crystal relief observation, especially when small amounts of bacteria were available.
Journal of Microscopy, Vol. 173, Pt. 1, January 1994, pp. 9--25.
Development, quantitative performance and applications of a parallel electron energy-loss spectrum imaging system
by G. BOTTON* & G. L'ESPERANCE,
Centre for Characterization and Microscopy of Materials, Departement de Metallurgie et Genie des Materiaux, Ecole Polytechnique de Montreal, C.P. 6079, Succ. ``A'' Montreal, (Quebec), Canada
Summary The development of a parallel electron energy-loss spectrum imaging system is presented. The analytical performance of the imaging technique was investigated and the system applied to materials science problems. The system, which allows acquisition and storage of a parallel electron energy-loss spectrum at each pixel of an image, was developed by interfacing a multichannel analyser and a microscope to a computer workstation. In the experimental conditions used for imaging, detection limits and quantification errors were large and varied as a function of spatial resolution and the range of chemical elements of interest in the image. Applications of this imaging technique in materials science showed that quantitative chemical information is provided by the system and that the use of relative thickness maps and detailed statistical analysis of the spectrum-image allowed an unbiased interpretation of the images. As energy-loss spectra are available after processing, spectroscopic information about the analysed material can be used to provide supplementary information.
Journal of Microscopy, Vol. 173, Pt. 1, January 1994, pp. 27--38.
Reflectance in situ hybridization (RISH): detection, by confocal reflectance laser microscopy, of gold-labelled riboprobes in breast cancer cell lines and histological specimens
by G. LINARES-CRUZ,* J. P. RIGAUT,"" J. VASSY,"" T. C. DE OLIVEIRA,"" P. DE CREMOUX,* B. OLOFSSON~~ & F. CALVO* ,
*Laboratoire de Pharmacologie Experimentale, Institut de Genetique Moleculaire, Hopital Saint-Louis, Universite Paris 7, 27 rue Juliette Dodu, F-75010 Paris, France. ""Laboratoire d'Analyse d'Images en Pathologie Cellulaire, U. 263 - I.N.S.E.R.M., Universite Paris 7, 2 place Jussieu, F-75251 Paris Cedex 05, France . ~~U. 248 - I.N.S.E.R.M., Faculte de Medecine Lariboisiere Saint-Louis, 10 avenue de Verdun, F-75010 Paris, France
Summary A method for reflectance in situ hybridization (RISH) is presented. The importance of the method is demonstrated by results obtained on cytological and histological breast cancer specimens. Scattering reflectance signals from 1-nm colloidal-gold particles after RNA/RNA in situ hybridization, using digoxigenin-labelled riboprobes, were detected by confocal scanning laser microscopy. The mRNA expression of two ras-related genes, rho B and rho C, was analysed in human histological breast cancer specimens and in human breast cancer cell lines. Horizontal (x, y) and vertical (z) optical sections after three-dimensional imaging were used for visualization. A marked heterogeneity (between individual cells and between specimens) was noted for the expression of the rho B gene, both in cytological and in histological samples. On the other hand, rho C was always expressed and showed no heterogeneity. This method allows the identification of several cellular constituents in an heterogeneous tissue structure, as demonstrated by the simultaneous detection of rho B (or rho C) by reflectance and of DNA, cytokeratin and/or vimentin by fluorescence.
Journal of Microscopy, Vol. 173, Pt. 1, January 1994, pp. 39--51.
Quantitative analysis of variable-angle total internal reflection fluorescence microscopy (VA-TIRFM) of cell/substrate contacts
by J. S. BURMEISTER, G. A. TRUSKEY & W. M. REICHERT* ,
Department of Biomedical Engineering, Duke University, Durham, NC 27708, U.S.A.
Summary Variable-angle total internal reflection fluorescence microscopy (VA-TIRFM) allows controlled variation of the illumination depth with the potential of measuring both membrane/substrate separation distances and sizes of focal contacts. VA-TIRFM images are collected from well-spread bovine aortic endothelial cells (BAEC) stained with a membrane-bound carbocyanine dye. Quantitative determination of absolute membrane/substrate separation distances and individual focal contact area are attempted using a simplified model of TIRFM optics. For angles slightly greater than the critical angle of 64 degrees, both the dorsal and ventral membranes were illuminated, while images excited above 66 degrees illuminated only focal contacts. Above 74 degrees the fluorescence of focal contacts was dominated by background noise. Direct application of the simplified optical model without accounting for background intensity was unsatisfactory. However, correction for background fluorescence and nonlinear regression of the untransformed data over the working range yielded focal contact separation distances of 24 (plus or minus 13) nm. Focal contact areas estimated by TIRFM (1.3 plus or minus 0.7 square micrometres) agreed closely with areas observed by immunofluorescence staining of vinculin (1.5 plus or minus 0.3 square micrometres).
Journal of Microscopy, Vol. 173, Pt. 1, January 1994, pp. 53--66.
Semiautomated methods for cancellous bone histomorphometry using a general-purpose video image analysis system
by W. E. HUFFER,* P. RUEGG,* J.-M. ZHU"" & R. B. LEPOFF* ,
*Department of Pathology, University of Colorado Health Sciences Center, 4200 East Ninth Avenue, Denver, Colorado, U.S.A.
Summary Semiautomated methods are used to measure elongated, curved and complex branching profiles and isolated perimeter segments in monochrome video images with a general-purpose analysis system. These methods are used to make the major primary measurements of bone histomorphometry. Accuracy and reproducibility of the image acquisition, processing and measurement system is documented by measuring a semicircular standard of known dimensions. Semiautomated applications of the Ar/Le method for measuring areas and perimeters, and calculating lengths and widths of osteoid seams, lengths of mineralization labels and mineral apposition rate, wall width, indirect measurements of eroded, osteoclastic and osteoblastic perimeters without tracing, and measurement of mineralized or total cancellous bone area and perimeter gave values comparable to measurements of the same parameters by tracing or grid counting techniques with equal or better reproducibility and much greater efficiency. Intraindividual variation in measuring multiple bone biopsies was comparable to that reported with current standard methods. Major sources of variability for semiautomated methods were image magnification and selection of profile edges by thresholding, and sources of variability for manual methods are image magnification, numbers of orthogonal intercepts, tracing speed and accuracy of the algorithm used to measure traced pixels. Semiautomated methods are accurate, reproducible and rapid methods suitable for bone histomorphometry.
Journal of Microscopy, Vol. 173, Pt. 1, January 1994, pp. 67--72.
Some remarks on the accuracy of surface area estimation using the spatial grid
by K. SANDAU* & U. HAHN , Institut fur Angewandte Mathematik und Statistik, Universitat Hohenheim, D-70593 Stuttgart, Germany
Summary A set of three line grids in three orthogonal directions is called a spatial grid. This spatial grid can be used for surface area estimation by counting the number of intersection points of a surface with the grid lines. If direction and localization of the spatial grid are suitably randomized, the expectation of this number is proportional to the surface area of interest. The method was especially developed for cases where the surface to be measured is embedded in a medium, which is the usual case in microscopical applications, and where a stack of serial optical sections of the surface is available. The paper presents an improvement of an earlier version of the counting rule for intersection points. Furthermore, if the direction of sectioning is not uniform random, a bias results. This bias is calculated for a disc as a perfectly anisotropic object. A generalization of the estimator is considered by introducing a weighted mean instead of the usual arithmetic mean. The variance due to the randomized direction is investigated depending on the weights, and the minimum of this variance is derived. The relationship between the covariogram and the variance of the surface area estimated with the spatial grid is considered.
Journal of Microscopy, Vol. 173, Pt. 1, January 1994, pp. 73--78.
Volume-weighted mean particle volume estimation using different measurement methods
by E. ARTACHO-PERULA & R. ROLDAN-VILLALOBOS,
Department of Morphological Sciences, School of Medicine, University of Cordoba, Avda. Menendez Pidal s/n, 14071 Cordoba, Spain
Summary The use of the `point-sampled intercepts' method on histopathological material is evaluated for the main purpose of comparing different methods of intercept length measurements. The volume-weighted mean nuclear volume of the carcinoma of the ampulla of Vater is calculated using three methods for measuring intercept lengths: a semiautomatic image analyser, an equidistant ruler and a logarithmic ruler. Rulers of several classes and lengths are used, and the results of the volume-weighted mean nuclear volume estimations are compared. The equidistant ruler is more accurate than the logarithmic ruler. With a greater number of ruler classes and with adjustment of ruler length to the greatest nuclear intercept lengths, the systematic deviation from the true value of the volume-weighted mean nuclear volume is smaller. The volume-weighted mean nuclear volume parameter has great power to differentiate intestinal carcinoma from normal tissue.
Journal of Microscopy, Vol. 173, Pt. 1, January 1994, pp. 79--82.
A simple calibration for routine section thickness measurements using current density ratios
by Y. M. HENG,* F. P. OTTENSMEYER,"" A. L. ARSENAULT* & G. T. SIMON*,
*Electron Microscopy Facility, McMaster University Medical Centre, 1200 Main St. West, Hamilton, Ontario L8N 3Z5, Canada
Summary A method to calibrate current density ratios for the determination of specimen thickness is presented. This method uses a tilt series from a single noncrystalline specimen to create different thicknesses; these are used to generate data points to establish the relationship between specimen thickness and current density ratio. The actual specimen thickness at 08 tilt was determined to an accuracy of 5nm by a parallax method. From the calibration curves obtained, we observed that the current density ratio was sensitive to relative thickness changes on the same section of less than 1nm when a 50-micrometre objective aperture was used.
Journal of Microscopy, Vol. 173, Pt. 1, January 1994, pp. 83--86.
A simple filter system for processing small or transparent specimens
by B. CRIBB & J. ZHU ,
Centre for Microscopy and Microanalysis, University of Queensland, Brisbane, Qld 4072, Australia
Summary A novel method of attaching a fine mesh filter to the end of a disposable plastic pipette is described. Such a pipette filter can be used to exclude specimen uptake during specimen preparation procedures, particularly when processing small or transparent materials. The pipette filter-tip does not interfere with fluid exchange and is non-reactive with normal processing fluids.
THE JOURNAL OF MICROSCOPY IS AVAILABLE AT A REDUCED RATE FOR MEMBERS OF THE ROYAL MICROSCOPICAL SOCIETY, THE MICROSCOPY SOCIETY OF AMERICA AND THE INTERNATIONAL SOCIETY FOR STEREOLOGY. PLEASE CONTACT THE SECRETARY OF EACH SOCIETY FOR DETAILS.
INSTRUCTIONS FOR AUTHORS AND DISK SUBMISSION FORMS CAN BE OBTAINED FROM DR GILLIAN WILSON, THE JOURNAL OF MICROSCOPY, 37/38 ST CLEMENTS, OXFORD OX4 1AJ, UNITED KINGDOM. TELEPHONE +44 (0)865 248768, FAX +44 (0)865 791237, EMAIL RMS-at-UK.AC.OX.VAX.
I would like to thank everyone for their help in my decalcification problem. Several techniques were suggested and I am sure one of them will work for my needs. Again, many thanks!
Subject: Time:2:31 PM OFFICE MEMO TEM: Mag Specs Date:1/4/94 TEM work on steel specimens can be very difficult, because they are almost always magnetized. You may be able to reduce the magnetic inhomogenieties a bit by passing the specimens through a strong AC field, a process industrially known as 'degaussing'. I believe degaussing coils can be purchased from most electronics supply stores that deal in the television market. In use, you insert the specimen into the center of the coil and withdraw it very slowly, with the AC current flowing through the coil.
Besides doing what Robert Keller suggests, you should note the objective lens current for a non-magnetic sample and set the lens to the same value. Then you can bring the specimen to the eucentric height by noting when the image is at the minimum contrast condition.
Russell Cook Electron Microscopy Center for Materials Research Argonne Natonal Laboratory Argonne, IL
Automatic print processing is a great benefit whether or not you also have an automatic exposure system. Indeed, we had a Kodak Royalprinter for many years before we aquired our LogEtronics (now Egoltronics) EM55 automatic dodging enlarger. Even now some of our users (not many) prefer to use our old Durst enlarger although they have only two simple aids to exposure determination: a Kodak Projection Print Scale (cat. no. 1557248) which cost about $10 and an Ilford EM10 Exposure Monitor which cost about $25.
Russell Cook Electron Microscopy Center for Materials Research Argonne Natonal Laboratory Argonne, IL
Subject: Time: 2:17 PM OFFICE MEMO TEM Analysis of Diffusion Couples Date: 1/4/94 We have considered using a TEM to study diffusion couples, but preparation of thin foils is a major stumbling block. Is anyone familiar with a possible technique that could leave the diffusion structure intact at the interface between two unlike materials in a diffusion couple? Is anyone aware of reported work relating to TEM analysis of diffusion couples?
There were several articles/abstracts in the Proceedings of the XIIth International Congress for Electron Microscopy (1990 EMSA Proceedings) on this subject, mine among them. If memory serves me, there are articles in the EMSA or MSA proceedings for years prior to and after 1990. My primary sample preparation technique, and that of many otherr researchers, is to ion-mill cross-sectioned samples.
Russell Cook Electron Microscopy Center for Materials Research Argonne Natonal Laboratory Argonne, IL
Eaton Publishing invites submission of papers for peer review in consideration for publication in the new journal "Cell Vision - Journal of Analytical Morphology." The first issue of Cell Vision is scheduled for publication in May/June 1994.
Cell Vision is edited for those scientists and physicians that analyze morphology as a means of diagnosis or research. It is also intended for those who are interested in advances in immunocytochemistry, confocal microscopy, image analysis and more recent developments such as in situ polymerase chain reaction and probe scanning microsopy.
Cell Vision focuses on these novel analytical methods in morphology and their applications in biomedical research and diagnostics. Developments reported in this journal will benefit any scientist who visualizes and analyzes chemical components against the background of tissue structure.
Cell Vision will have an international circulation and will publish articles contributed by multinational authors. All articles will be rigorously peer reviewed and promise to be of very high quality. The international Editorial Board of Cell Vision, led by Dr. Jiang Gu of the Deborah Research Institute, includes many top scientists in modern morphology.
General information about Cell Vision (including instructions for authors and subscription information) is available by contacting Eaton Publishing, 154 East Central Street, Suite 201, Natick, MA 01760. You may also fax your requests to 508-655-9910 or submit electronic mail requests to internet address: fweaton-at-biotechnet.com.
Thanks for your comments. I'm headed off to look at my samples this afternoon and see how the suggestions work. For those who are wondering, I prepare my samples by cutting with a wire saw to 100microns and slowly (as possible) jet polish with perchloric until there is a hole.
We have just taken on a project which involves cutting "thin" sections of 35 um fenestrated glass beads. I am using tungsten coated glass knives broken via the "balanced-break" method with , as expected, less than desireable results. Has anyone had any experience with anything like this? Incidentaly, the beads are fixed and embedded in Spurr's resin. Any ideas for better sections would be appreciated.
Subject: Time:1:13 PM OFFICE MEMO Re} EM Atlas Date:1/5/94 {I am looking for EM atlases (atli?) for both "normal" and pathologic {tissues. {I work almost exclusively with mouse tissue, but any good atlas should {be enough of a guide. I'd appreciate any infomation or suggestions. Margaret, You did not mention the type of tissue you are looking at. If you are interested in nervous system tissue, the definitive "atlas" is, The Fine Structure of the Nervous System, by Peters, Palay and Webster. The most recent edition (3rd) is published by Oxford. Mike Mike_Schwartz-at-qm.yale.edu
} Date: Wed, 5 Jan 94 12:15:01 EST } From: meh-at-jax.org (Margaret E. Hogan) } To: microscopy-at-anlemc.msd.anl.gov } Subject: EM Atlas
} I am looking for EM atlases (atli?) for both "normal" and pathologic tissues. } I work almost exclusively with mouse tissue, but any good atlas should be } enough of a guide. I'd appreciate any infomation or suggestions. Thanks!!!! } } Peggy Hogan } The Jackson Laboratory } meh-at-aretha.jax.org
Another atlas, not the usual normal or pathological atlas, but very useful: Artifacts in Biological Electron Microscopy R.E.F. Crang and K.L. Klomparens, eds. Plenum Press, NY 1988
Organisers: Dr A Entwistle Dr C V Howard Dr H Gundlach
The Digital Imaging Light Microscopy Summer School is aimed primarily at scientists in biological and related disciplines and will comprise of a basic introduction, followed by a selection of modules.
-------------------------------------------------------------- Monday and Tuesday
Common Core - Introduction to Light Microscopy. This will consist of lectures, demonstrations and practical work on the following:
The History of microscopy; Introduction to microscopy; Limitations of the eye; Resolution, Contrast, Magnification; Refraction and lenses, geometrical optics, conjugate planes; Aperture; Illumination of the specimen in transmitted and reflected light; Lens aberrations and their correction; the choice of optical components; Diffraction and its consequences for the microscope image; Generation of contrast; Photomicrography.
Modules - The following five options are available:
Comprehensive Digital Light Microscopy - Stereology and digital imaging, digital image collection, digital image processing, digital image display and digital image storage.
Confocal Microscopy (students must bring their own specimens for study) - Fluorescent staining of samples, imaging of samples (fluorescence, reflectance and DIC techniques), visualization of 2-D and -D data sets and aspects of confocal theory.
Advanced Fluorescence Microscopy - Fluorescent staining of samples, ion ratioing methods, fluorescence decay-time measurements, fluorescence confocal microscopy.
Stereology and Digital Light Microscopy - Basic concepts of systematic random (ie unbiased) sampling in 3-D from histological material, efficient design-based stereological methods for estimating number, surface, length and volume (both manual methods and those employing image analysis systems will be covered) and methods of particle sizing in 3-D.
Techniques in Digital Light Microscopy - Introduction to stereology, introduction to confocal microscopy and introduction to digital imaging and processing.
FOR FURTHER DETAILS CONTACT THE ROYAL MICROSCOPICAL SOCIETY, 37/38 ST CLEMENTS, OXFORD OX4 1AJ, UNITED KINGDOM. TELEPHONE +44 (0)865 248768, FAX +44 (0)865 791237, EMAIL: RMS-at-UK.AC.OX.VAX.
I'm interested in the morphology of Panthera Pardus epidermis. Does anyone know if there is something unusual about the epithelial cells of this animal skin?
============================================================================= Francisco Javier Hernandez Blazquez | Universidade de Sao Paulo - Faculdade de | e-mail fjhblazq-at-fox.cce.usp.br Zootecnia e Engenharia de Alimentos | Voice: 55 195 616122 r. 265 Departamento de Ciencias Basicas/Histologia| Av. Duque de Caxias Norte, 225 CP 23 | Fax: 55 195 611689 CEP 13630-000 Pirassununga (Sao Paulo) | BRAZIL | ==============================================================================
Charles Bradley at Argonne National Laboratory has been sectioning very small bits of radioactive waste glasses embedded in resin. I believe the size of the glass pieces are about 40 microns or so. I am not sure of the resin he has been using, however. He has been using a diamond knife to get TEM sections from a Reichert-Jung Ultracut E.
Russell Cook Electron Microscopy Center for Materials Research Argonne Natonal Laboratory Argonne, IL
I have come across a structure, many times, that I have identified as a perichromatin granule. A dense body of about 30-40 nm surrounded by a transparent halo an usually associated with dense chromatin. I think this structure was first mentioned by Bernhard 20 or more years ago. Does any one know if this structure has been further characterized? Any leads on this would be greatly appreciated.
A paper by Bottke may be of interest: Bottke W (1976). Chromosome-associated paracrystalline nuclear inclusions in the spermatocytes of a pulmonate snail, Planorbarius corneus L. Chromosoma (Berl.) 55: 273-287.
On Thu, 6 Jan 1994, Greg Erdos ICBR EM Core Lab University of Florida wrote:
} Cell Biologists: } } I have come across a structure, many times, that I have identified } as a perichromatin granule. A dense body of about 30-40 nm surrounded by a } transparent halo an usually associated with dense chromatin. I think this } structure was first mentioned by Bernhard 20 or more years ago. Does any } one know if this structure has been further characterized? } Any leads on this would be greatly appreciated. } } ***************************** } * Greg Erdos * } * Director, ICBR EMCL * } * University of Florida * } * Gainesville, FL 32611 * } * gwerdos-at-gnv.ifas.ufl.edu * } * 904-392-1295 * } ***************************** }
With encouragement from a number of you folks out there, I tried sectioning these thing with an old diamond knive. Much to my amazement, it worked beautifully. As a biologist, I never would have thought that this was possible. We have good sections with minimal obvious damage to the knife. We have saved a great deal of time and many headaches. Thanks to all who responded and HOORAH for the system.
We are looking for a used but recent vintage electron microprobe to replace a very old ARL system. Preferences are for a 4-channel system with EDS, something like a Cameca SX-50. We are aware of Don Lesher's operation in northern Ohio and have talked with him about acquiring a rebuilt ARL-SEMQ. This may be our eventual route, but if there is a good recent vintage machine sitting out there somewhere we would be very interested in knowing about it. Thanks. Warren D. Huff Dept. of Geology University of Cincinnati Cincinnati, OH 45221-0013 phone (513) 556-3731 fax (513) 556-6931 e-mail huff-at-ucbeh.san.uc.edu
Due to the continuing recession in California and especially the hardships the UC system is undergoing we are forced to begin a recharge for our microscopy services. I would very much like to know what current recharges are for biological TEM use and prep services. If anyone has a freeze fracture device (we have a Balzers BAF 400T) what are the going rates? How about for SEM prep?
Thank you in advance. Rick A. Harris Electron Microscopy Evolution and Ecology Univ. of Calif. Davis, CA 916 752 2914
None Re: Recent request for help on volume calculations -
"Our laboratory has expressed interest in calculating volumes on microscopic data and I am looking for software that can perform the following tasks:
2: 3-D reconstruction (including EM image alignment) 5: Wire-frame generation and simple volume rendering 3: Volume calculation 1: Run on a Mac or PC 4: File I/O from a Mac or PC format (preferrably using MacDraw objects of known dimensions)
The non-proprietary file format support is neccessary to allow cross-platform data manipulation. My U*NIX access is limited, so ability to un on a PC is important.
I keep reading how people did 3-D reconstruction, but their programs are hard to find. Any referrals to sources of this software is appreciated. It WOULD be neat if it was available in C by anonymous FTP, but perhaps this is too narrow a requirement. Any leads would help."
Reply - There are simple methods for estimating volume densities that do not require complicated computer systems or sophisticated software. If your aims are to estimate the volumes of subcellular structures and you can directly measure at least one reference volume then you might like to try cross latice overlays as described by Weibel as far back as 1979 (Stereological methods vol 1. practical methods for biological morphometry Academic Press NY). More recent reviews can be found in APMIS (Gundersen et al 1988 vol 96 379-394), J. Microsc (Cruz-Orive 1982 vol 125 89-102) Am. J. Physiol (Cruz-Orive & Weibel 1990) vol 258 148-156) and TICB (Luquoc 1993 some time this fall). Another source is a recently published book where the final chapter covers all these methods (Griffiths 1993 Fine Structure Immunocytochemistry, Springer Verlag, Heidelberg). Best of all is to take a course on stereology (there is one in Banff, Canada in May 1994 and one here at Yale in August 1994). If you still want to do 3-D reconstructions, which will only give you information on the structures you reconstruct, then the best software I have seen up to now are the VoxelView programs. We run them on a Silicon Graphics workstation which never seems to have enough memory. These programs allow you to reconstruct sections, rotate and manipulate the images as well as measure the parameters of the reconstructed structures.
Immunocytochemistry and Cryosections Practical Course 22 - 27 August 1994. An intensive practical course mixed with theoretical sessions where you can learn how to produce cryosections as well as immunolabeling, colloidal gold production and much more.
Additional we are offerring a three day practical workshop on Stereological methods. This will be on 18 -20 August 1994, prior to the cryosectioning course.
A team of instructors headed by Hans Gundersen will take you through the theoretical and practical details of modern stereological methods. These will include volume and surface densities, the fractionator, the nucleator, the disector and much much more. Address for further details on both courses;
Paul Webster, Department of Cell Biology, Yale School of Medicine, 333 Cedar Street, New Haven, CT 06510.
Re} Stereology Re: Recent request for help on volume calculations -
"Our laboratory has expressed interest in calculating volumes on microscopic data and I am looking for software that can perform the following tasks:
2: 3-D reconstruction (including EM image alignment) 5: Wire-frame generation and simple volume rendering 3: Volume calculation 1: Run on a Mac or PC 4: File I/O from a Mac or PC format (preferrably using MacDraw objects of known dimensions)
The non-proprietary file format support is neccessary to allow cross-platform data manipulation. My U*NIX access is limited, so ability to un on a PC is important.
I keep reading how people did 3-D reconstruction, but their programs are hard to find. Any referrals to sources of this software is appreciated. It WOULD be neat if it was available in C by anonymous FTP, but perhaps this is too narrow a requirement. Any leads would help."
Reply - There are simple methods for estimating volume densities that do not require complicated computer systems or sophisticated software. If your aims are to estimate the volumes of subcellular structures and you can directly measure at least one reference volume then you might like to try cross latice overlays as described by Weibel as far back as 1979 (Stereological methods vol 1. practical methods for biological morphometry Academic Press NY). More recent reviews can be found in APMIS (Gundersen et al 1988 vol 96 379-394), J. Microsc (Cruz-Orive 1982 vol 125 89-102) Am. J. Physiol (Cruz-Orive & Weibel 1990) vol 258 148-156) and TICB (Luquoc 1993 some time this fall). Another source is a recently published book where the final chapter covers all these methods (Griffiths 1993 Fine Structure Immunocytochemistry, Springer Verlag, Heidelberg). Best of all is to take a course on stereology (there is one in Banff, Canada in May 1994 and one here at Yale in August 1994). If you still want to do 3-D reconstructions, which will only give you information on the structures you reconstruct, then the best software I have seen up to now are the VoxelView programs. We run them on a Silicon Graphic workstation which never seems to have enough memory. These programs allow you to reconstruct sections, rotate and manipulate the images as well as measure the parameters of the reconstructed structures.
There is a tutorial on quantitative morphometry written by Dr. Robert Bolender, Dept. Biological Structure at Univ. Washington. It also provides templates for point and intersect counts and for QM computations. It is available from the Health Sciences Center for Educational Resources, University of Washington SB-56, Seattle, Washington, 98195 (206) 685-1156.
Dr. Bolender teaches a course on QM in even years.
Glen MacDonald Hearing Development Laboratories RL-30 University of Washington Seattle, WA 98195 (206)543-8360 glenmac-at-u.washington.edu
Subscribers who might find it difficult to attend the stereology courses in Banff or Yale should note that the RMS will be running a Digital Imaging and Stereology course at Liverpool University, UK, in July 1994, coordinated by the current President of the International Society for Stereology, Dr Vyvyan Howard. Contact RMS-at-UK.AC.OX.VAX with your full name and address to obtain details!
From Paul Sheppard, Tree-Ring Lab, Univ. of Arizona
To microscopy forum members:
I am having slight, but definite, difficulty in attaining repeatable focus of my binocular common main objective microscope. In my attempt to apply image- analysis techniques to tree-ring science, my focus problem has led to systemat- ic differences in values as measured by different technicians. I am amazed at how little the focus differences must be before we see differences in our data. Can anyone suggest ways to ensure repeatable focus?
I have inquired into adding on autofocus hard- and software, but my first estimate on that was $7,000, which is prohibitive at this time. I have also heard about a dual-light focus aid, where two spot beams are projected onto the subject to intersect exactly at the focal distance of the objective lens; if the subject is above or below that distance, then the spot will be elongated or even split into two. Has anyone tried this? I have also tried focussing at high magnification and then working at my usual lower magnification. This re- quires parfocal optics, which I have, but this process is cumbersome and other- wise prone to error.
Thanks in advance for any and all suggestions,
Paul Sheppard Laboratory of Tree-Ring Research University of Arizona GRAD12-at-CCIT.ARIZONA.EDU
My work is in the materials sciences where for reflected light microscopy specien preparation usually begins with a grinding and polishing sequence in order to obtain an optically smooth surface. I am now faced with a material I do not wish to grind/polish, I passed by a reference that suggested that if the specimen surface is not optically smooth I can use glass coverslips and an index matching fluid - this sounds vaguely familar from highschool biology. Is there anyone out there with experience in Light Microscopy who could point toward the correct products and describe for me some of the considerations involved.
} Rick Harris asked about charges for EM facilities..... } --------------------------------------------------- } } Sandy Silvers of the South Eastern EM Society has a fairly } comprehensive database on EM facilities in the US. You } can contact her at
** some stuff deleted **
I just got off the phone with Sandy Silvers and told her that, since she doesn't currently have Internet access, I'd pass this information along. She has moved from Florida. Her current address and phone number are:
Sandra Silvers EM Complex USDA, ARS, RRC PO Box 5677 Athens GA 30613-6199 (706) 546-3471
Her database has listings, which include contact people, for about 200 facilities. Information about usage charges is included for most of them. She is looking for more input and will send a questionaire on request. Printed copies of the database are available for $8 (US). She does all this personally now, so needs to recoup printing costs.
She's intrigued with the possibility of having the information available more widely, perhaps through FTP.
John chandler-at-lamar.ColoState.EDU Fort Collins, CO
Subject: Time:4:14 PM OFFICE MEMO LM???-Reply Date:1/11/94 I do not know the nature of your specimens, but I seriously dobut that you would gain much useful information by using a cover slide and oil to overcome the usual polishing process. Polishing is usually necessary because optical microscopes have such limited depth of field (Typically about 10 microns for a 10X objective, 1 micron for a 40X obj, and 0.1 or 0.2 microns for a 100X obj) that they are not at all suited for looking at rough surfaces. Placing a layer of oil and a cover glass over a rough surface will not solve this problem. Furthermore, the objective lenses of metallographic microscopes are not designed to work through cover glasses - you'll probably need to go to the biology or mineralogy dept to find a microscope that is. I would suggest that you try examining the specimen with a stereo binocular microscope as a start. A good instrument of this kind should give magnifications up to about 75X. If that is not sufficient, then the next step would be to try SEM - although contrast may be a problem, depending on the nature of the specimens. microscope will not work with a cover glass,
The previously posted information about Sandy Silvers address was incorrect. She has since moved, her new address is apparently.... Thanks to L. Melsen for the correction....
Rick Harris asked about charges for EM facilities..... ---------------------------------------------------
Sandy Silvers of the South Eastern EM Society has a fairly comprehensive database on EM facilities in the US. You can contact her at
SANDY H. SILVERS EM COMPLEX RUSSELL RESEARCH CENTERS USDA, BOX 5677 ATHENS, GA 30613-6199 (706) 546 3471 FAX (706) 546 3452
Subject: Time:5:15 PM OFFICE MEMO Stereographic Calcs Date:1/11/94 We have obtained optical goniometric measurements of angles between the faces of about 75 macro crystals. These angles (commonly called phi and rho) were measured relative to the axes of the optical goniometer. Now, we want to convert them to angles that are relative to principal planes of the crystals. Does anyone have , or know of, a readily available computer program for doing this?
We are working on a project that requires us to shadow myosin with Pt and C. We are trying to see the myosin heads and another conjugated molecule. We are using our Balzers 400 to shadow at 15 to 25 degrees for the Pt and then at 90 degrees for the C. We can find the molecules but they are extremely faint. We have used anywhere from 5 to 15 angstroms of Pt. Best results were with higher angles and thinner coats. Then we increase exposure in the TEM to 3 seconds to bump the contrast and shift the s/n ratio. Has anyone a suggestion for making the myosin more visible? The micrographs have little contrast between the molecule and the substrate.
Rick A. Harris Electron Microscopy Evolution and Ecology Univ. of Calif. Davis, CA
Excerpts from mail: 11-Jan-94 LM - ??? by Peter Joyce-at-utxvms.cc.ut } My work is in the materials sciences where for reflected light microscopy } specien preparation usually begins with a grinding and polishing sequence } in order to obtain an optically smooth surface. I am now faced with a } material I do not wish to grind/polish, I passed by a reference that } suggested that if the specimen surface is not optically smooth I can use } glass coverslips and an index matching fluid - this sounds vaguely familar } If I understand your question, I don't think index matching will help. Reflections occur where refractive index changes, if you match the refractive index of your specimen you not see much reflection. I am not a materials scientist, but biologists use reflected light microscopy to generate contrast by interference of the first surface reflection (usually the coverslip/medium interface) with the second surface reflection (usually the medium/specimen interface) in order to see regions where the specimen makes close contact with the coverslip. If your application is based at all on similar effects, index matching will at least attenuate one of the reflections.
As part of a research project, I am looking for -any and all- references that pertain to the staining of mineral species for their characteriza- tion and to make certain minerals more distinguishable under the optical (polarizing) microscope.
I greatly appreciate all info and help in this endeaver.
Thanks, Rob
X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X X Rob Tayloe X MSM Spelunkers Club X X Metallographic Lab. X Missouri Speleological Survey /-v-\ \-v-/ X X Rolla Research Center X Bat Conservation International X X U.S. Bureau of Mines X Missouri Cave & Karst Conservancy \-v-/ X X tayloe-at-rorc.usbm.gov X National Speleological Society #32993 /-v-\ X X (314) 364-3169 x247 X American Cave Conservation Association X X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X
I am wondering if anyone out there has tried incorporating the new Mac AV computers with their video systems? I've got the money for the Mac, but I need to know how the software is for image analysis, and how the final product actually looks.
If anyone has done an EM video with the Mac I would like to see it.
Thanks for the help...Go Giants...long live the VW!
John L. Grazul BBR EM Facility Rutgers University Box 1059 Piscataway, NJ 08854
On the NIH-Image mailing list, many, many postings have gone through with the general concensus being that the AV input on the Macs was not intended for scientific quality. There are a number of high-quality frame grabbers for 640 x 480 pixel pictures (e.g., Scion LG-3 at 301-695-7870, Data Translation QuickCapture at 508-481-3700, Perceptics at 615-966-9200, others) as well as some higher-end systems for cooled CCDs, etc. There are also some slow-scan TEM interfaces for the Mac (Gatan, 4pi Analysis at 919-489-1757 for STEM, others). There has been considerably less discussion about the video output. If this is the issue, someone else should comment.
As for image analysis software, there are a number of packages ranging from classical microscopic image analysis to part inspection on assembly lines. My choices are NIH Image (free from NIH via anonymous FTP to zippy.nimh.nih.gov) for workhorse viewing and automation as well as "simple" particle analysis and PrismView (marketed by Signal Analytics 703-281-3277) for high-end work. NIH Image is quick to learn, readily extensible and is extremely popular with its infinite return-on-investment! There are other systems as well.
These vendor lists are NOT exhaustive. If you can download NIH Image, appendices B,C, and D are a little more exhaustive for Mac imaging vendors.
Bill ========================== Bill Heeschen / Analytical Sciences - Materials Characterization 1897-D Building / The Dow Chemical Company Midland, MI 48667 U.S.A. phone: (517)636-4005 fax: (517)636-5453 Email: waheeschen-at-dow.com ==========================
} } As for image analysis software, there are a number of packages ranging from } classical microscopic image analysis to part inspection on assembly lines. My } choices are NIH Image (free from NIH via anonymous FTP to zippy.nimh.nih.gov) } for workhorse viewing and automation as well as "simple" particle analysis and } PrismView (marketed by Signal Analytics 703-281-3277) for high-end work. NIH } Image is quick to learn, readily extensible and is extremely popular with its } infinite return-on-investment! There are other systems as well. } } These vendor lists are NOT exhaustive. If you can download NIH Image, } appendices B,C, and D are a little more exhaustive for Mac imaging vendors. } } Bill
Another note regarding easy info on NIH Image, subscribe to the mailing list located on nih-image-at-soils.umn.edu. They're alll the time fielding most any question you can think of, including questions regrading PrismView. If it's too much to subscribe, post your question anyway and have replies sent directly to you. Good luck.
I was wondering if there was a software package similar to NIH-Image that will run on the IBM or Silicon Graphics workstation that one can get through ftp.
I need a small sample of Ti203 for some microscopy work I'm doing. It's only a really small amount needed so If someone has some that they can give me I would be most appreciative, failing that does anyone know of a supplier?
Thanks David
-- ---------------------------------------------------------------------- David Bell |E-mail: dcb-at-electron.ph.unimelb.edu.au School of Physics |Phone : +61 3 344 5451 The University of Melbourne |Fax : +61 3 344 4783 Parkville, Victoria, AUST, 3052 |
Here is a supplier for Ti2O3, otherwise known as titanium (III) oxide: AESAR/Johnson Matthey 30 Bond Street P.O. Box 8247 Ward Hill, MA 01835-0747 (800) 343-1990 (508) 521-6300. You should be able to get 50 grams for about $50.
J. Ester asked } I was wondering if there was a software package similar to NIH-Image } that will run on the IBM or Silicon Graphics workstation that one can } get through ftp
There is no equivalent for NIH Image that will run on the PC. The closest thing (and it's not close) is NCSA Image fro the National Center for Supercomputing Applications at the Univ. of Ill. It is available via FTP. Their address is "ftp.ncsa.uiuc.edu". NIH Image as I understand from Wayne Rasband (the author at NIH) will be ported to the PowerPC when he gets a unit. So those of you who are tied to PC will be able to run as long as you run the machine in it's Mac Compatible Mode instead of the PC Compatible Mode.
Department of Anatomy and Neurobiology Colorado State University Fort Collins, CO
July 11-15, 1994
The CSU Electron Microscopy Center in the Department of Anatomy and Neurobiology offers an intensive course in freeze-fracture and freeze-etch techniques for research scientists and senior technicians.
Basic and advanced freeze-fracture and freeze-etch techniques will be taught in five days of concentrated lectures and laboratory sessions (12-15 hr/day). Advanced techniques will include sequential confocal mapping/freeze-fracture examination of identified cells in tissue slices. This course will prepare research scientists and laboratory technicians to use freeze-fracture techniques in cell biology research. No previous freeze-fracture experience is necessary, but the individual must be proficient in transmission electron microscopy.
Participants will prepare high-resolution replicas of their own specimens, become proficient at interpreting the resulting images, and learn to prepare and label their own stereoscopic micrographs. Faculty and participants will discuss the physical and chemical bases for interpreting freeze-etch replicas, the major sources of specimen preparation artifacts, and the unique advantages and limitations of freeze-fracture and freeze-etch techniques. Methods for reducing specimen contamination during the fracturing and replication processes, as well as techniques for identifying, fracturing, and mapping individual cells in tissue slices, will be described.
This course is organized by Drs. John Rash, John Walrond, Robert Lee and John Chandler in collaboration with major instrument manufacturers.
Freeze-fracture/freeze-etch equipment used in the course includes: Balzers BAF-301 and BAF- 400, JEOL JFD-9010-CR and additional rapid freeze and freeze-etch devices supplied by RMC, Inc. and Bal-Tec, Inc. Molecular Dynamics multi-probe 2001 confocal laser scanning microscope will be used.
Registration fee of $1250 includes textbook, laboratory instruction manual, all necessary supplies and implements, noon meals, refreshments, EM negatives and prints, and unlimited use of equipment during the course.
Information concerning this course, hotel accommodations and travel arrangements may be obtained from:
Eileen Diepenbrock Colorado State University Department of Anatomy and Neurobiology Fort Collins, CO 80523 (303) 491-5847 ediepenbrock-at-vines.colostate.edu
Registration closes May 15, 1994
Course registration will be for a minimum of 10 and maximum of 12 participants.
John chandler-at-lamar.ColoState.EDU Fort Collins, CO
I've had multiple requests lately for permission to post resume's and job "hunting notices" on this listserver. As we set forth early in the setup of this discussion/information forum that type of posting is not appropriate here. However, I realize that with the current budget situations across the country & world there will be a significant number of highly talented individuals looking for positions in the near term. Allow me then to remind all of you that should you have acess to information concerning research/teaching positions this type of BULLETIN/ANNOUNCEMENT is allowed on the Mailserver and I would encourage you to post it. If you are not sure about any posting feel free to Email it directly to me and I will review it's appropriateness.
Individuals interested in posting (SHORT) electronic resume's are welcome to access the ANLEMC/MSA electronic bulletin board. It can be reached via INTERNET/TELNET link at the address: ANLEMC.MSD.ANL.GOV, login with the username EMCBBS and password EMCBBS then follow directions on the screen. Please note that there is only a single connection from INTERNET to this BBS line and that the line may be in use. Pay attention to any messages on your terminal when you login this will clue you in to what is happening....
I received a copy of your response to John L. Grazul regarding digital TEM. We are using a system consisting of a Gatan wide angle video camera (model 673mkIII ), a Dage video processor (DSP200), Quick Capture board and a MacII running NIH-Image. The combination yields 640x480 images directly from the microscope. With the frame processor we can average frames (2,4,8,16,32) and adjust the gain and offset of the video signal so the Mac receives a full range signal. I have written some macros for Image to permit rapid specimen ID, magnification setting and storage. Folders of images are then sent over the network to the pathologist's desk.
We have found that the images are sufficient for diagnostic purposes and have begun to shift to electronic imaging.
The images are NOT the quality of film, but the cost for the system is low (NIH Image is free and VERY good). My concern was (and is) that with a slow scan system the increased cost doesn't really get you that much more in terms of resolution (maybe twice the resolution for lots more money). The microscope has such excess magnification that it is much cheaper to acquire multiple images at higher magnification (and hence greater resolution) than it is to purchase a slow scan camera and the associated software drivers. My thought is that if you want film-like resolution you should take micrographs and either print them or digitize the negative with a flatbed or drum scanner for image analysis. If all you want to do is some image analysis, it is entirely possible that a 640x480 image will be sufficient.
Feel free to forward this to Grazul and others on whatever mail list or news group you are on.
I would be interested to learn how you are using your system.
Charles Daghlian Rippel E. M. Facility Dartmouth College Hanover, NH 03755 603-650-1337
The list administrator is John Ladwig (jladwig-at-soils.umn.edu), but don't bug him until you've tried the above instructions! 8-)
As many of you list participants are aware, there is movement afoot to set up a NewsGroup which will be a superset of both the nih-image and microscopy lists. As this develops, there will be postings to both lists describing how/when this happens.
Bill ========================== Bill Heeschen / Analytical Sciences - Materials Characterization 1897-D Building / The Dow Chemical Company Midland, MI 48667 U.S.A. phone: (517)636-4005 fax: (517)636-5453 Email: waheeschen-at-dow.com ==========================
* SC3 Ion Projection Lithography Instructor: John C. Wolfe, Univ. of Houston
Resists -------
* SC4 Introduction to Microlithography, Resist Materials and Processing Instructors: Larry F. Thompson, AT&T Bell Labs.; Murrae J. Bowden, Bell Communications Research, Inc.; C. G. Willson, Univ. of Texas/Austin
* SC5 Optical Lithography Modeling Instructors: Chris A. Mack, FINLE Technologies, Inc.; Andrew R. Neureuther, Univ. of California/Berkeley
* SC6 Resist Thickness, Bake, Exposure, and Development Control Instructor: W. Tom Batchelder, Semiconductor Systems, Inc.
* SC7 Resists for Deep-UV Lithography Instructor: C. Grant Willson, Univ. of Texas/Austin
* SC8 Diazonaphthoquinone-Based Resists Instructor: Ralph Dammel, Hoechst Celanese Corp.
Metrology & Process Control ---------------------------
* SC9 IC Critical Dimension Measurement System Instructors: Sadri Khalessi, Metrologix, Inc.; Kevin M. Monahan, Metrologix, Inc.
* SC10 Fundamentals and Pitfalls of Submicrometer Dimensional Metrology Instructor: Terrence E. Zavecz, TEA Systems Corp.
* SC11 The Physics and Simulation of Metrology Instruments Instructor: Mark P. Davidson, Spectel Co.
* SC14 The Use of the Low-Voltage SEM in IC Fabrication Instructor: Michael G. Rosenfield, IBM Thomas J. Watson Research Ctr.
Microlithography ----------------
* SC15 Advanced Topics in Optical Lithography Instructor: Chris A. Mack, FINLE Technologies, Inc.
* SC16 Fundamentals of Cameras and Projectors Instructor: Warren J. Smith, Kaiser Electro-Optics, Inc.
* SC17 Introduction to Optical Lithographic Tools Instructor: Timothy A. Brunner, IBM Thomas J. Watson Research Ctr.
* SC18 Plasma Etching Technology Instructor: Daniel L. Flamm, Univ. of California/Berkeley
* SC19 The Exposure-Defocus Tree and Its Uses in Understanding and Extending Optical Lithography Instructor: Burn J. Lin, Linnovation, Inc.
* SC20 Optimization Methods for Microlithographic Materials and Processes Instructor: Daniel J. Herr, Semiconductor Research Corp.
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Don Grimes of Microscopy Today has sent the enclosed message along to me for approval for posting on the Microscopy Mailserver. I do not have a problem with it as it is a general announcement meant as a service to Microscopists who will be looking for jobs.
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To: Membership
Thanks to many for your interest and comments regarding our newsletter. Several have inquired over the possibility of our assisting in their search for new employment. I have previously not done this as it does not quite meet my objective as "material of interest to a reasonable number of microscopists". However I would like to try to help. If you seek new employment and would like to provide me a summary of either "what" you are or "what" you are looking for, I will publish the summary in my next issue. I only ask that you keep the summary under around 60 words and I MUST have it no later than this coming Friday (21 Jan) to make the issue. And if you wish to keep your name "quiet", we can do the Box # thing - where a number rather than your name is listed, and I will forward any interests direct to you. I expect that my newsletter is received in 99+% of the microscopy labs in the U.S. but can not guarantee that it goes to the right person.
Due to the number of replies to the attached message here is more info. The QM2000 tutorial runs on DOS and Windows. Its development platform is due out for the Mac and Unix, so there may be ports to those operating systems. Dr. Bolender has released a quantitative morphometry dataabase for the nervous system, developed with Sybase, running on a Sparc.
A summer quarter course in quantitative morphometry will probably be taught here this summer.
For more information: Dr. Robert Bolender Dept. Biological Structure SM-20 University of Washington Seattle, WA 98195 rpb-at-u.washington.edu
******old message There is a tutorial on quantitative morphometry written by Dr. Robert Bolender, Dept. Biological Structure at Univ. Washington. It also provides templates for point and intersect counts and for QM computations. It is available from the Health Sciences Center for Educational Resources, University of Washington SB-56, Seattle, Washington, 98195 (206) 685-1156.
Dr. Bolender teaches a course on QM in even years.
Glen MacDonald Hearing Development Laboratories RL-30 University of Washington Seattle, WA 98195 (206)543-8360 glenmac-at-u.washington.edu
I will am about to start as a postdoctoral scientist working with Prof. Kirz at the State University of New York at Stony Brook/Brookhaven National Laboratory in the field of x-ray microscopy. I am very interested in your EM-course on Immunocytochemistry and Cryosectioning. Please let me know what I need in order to apply. Sincerely, Joerg Maser
The Department of Materials Science and Engineering at Lehigh University has an opening for a senior (associate/full) professor with expertise in electron microscopy and/or microanalysis (SEM, TEM, AEM, etc.). The successful candidate must have a proven record of research in the application of microscopy and/or microanalysis to the solution of materials problems and be able to conduct independent and cooperative research in MS&E. Ability to teach undergraduate and graduate courses in microscopy and materials is essential. Experience in running a microscopy facility will be an advantage. Position available September 1, 1994.
Curriculum vitae and the names of three references should be sent by May 15, 1994 to: Professor David B. Williams Chairman, Search Committee Dept. of Materials Science and Eng. Lehigh University 5 East Packer Avenue Bethlehem, PA 18015-3195
Lehigh University is an equal opportunnity employer and welcomes applications from all qualified candidates.
Thanks to B.Miner for forwarding the useful comments from P.Catinella and B.Edwards on RIE etching on microelectronic devices.
One other question that I would like to direct to them or other interested parties would be the requirement for installing a "scrubber" on the exhaust fumes from the RIE. We are working up a estimate for the possible total cost of purchasing/operating/maintaining a plasma etcher in our laborato- ry. The requirement for a "scrubber" on the exhaust has been mentioned as a possibile environmental requirement. I have spoken to some manufactures and they don't use scubbers due to low flow rates in their machines and small quantities of gas involved.
Thanks again.
Richard Sartore US Army Research LAboratory AMSRL-EP-RA Fort Monmouth, NJ 07703 RSARTORE-at-MONMOUTH-ETDL1.ARMY.MIL
To Membership, International (or local) readers who wish to receive a no cost copy of our newsletter "Microscopy Today" and are having trouble contacting me, as I am on Compuserve, are encouraged to do so by FAX. My number is (608)836-1969. And the several who have requested the newsletter but did not supply their addresses are encouraged to resent their requests.
We have a Nanoscope III here, and would like to study the images off-line on another pc. Does anyone know good image analysing & processing programs for the PC? Basic filtering and measurement functions (cross sections etc) are a must.
I've heard names like Global Lab Image, Mocha, Image Pro Plus. Any experiences on these? Where to take contact: companies, addresses, fax numbers?
Thanks in advance,
Markus Levlin
-- Markus Levlin Laboratory of Physics tel +358 0 451 3144 markus.levlin-at-hut.fi Helsinki University of Technology fax +358 0 451 3116 Otakaari 1 M, 02150 Espoo, Finland
Thanks to B.Miner for forwarding the useful comments from P.Catinella and B.Edwards on RIE etching microelectronic devices.
One other question that I would like to direct to them or other interested parties would be the requirement for installing a "scrubber" on the exhaust fumes from the RIE. We are working up a estimate for the possible total cost of purchasing/operating/maintaining a plasma etcher in our laborato- ry. The requirement for a "scrubber" on the exhaust has been mentioned as a possibile environmental requirement. I have spoken to some manufactures and they don't use scubbers due to low flow rates in their machines and small quantities of gas involved.
Thanks again.
Richard Sartore US Army Research LAboratory AMSRL-EP-RA Fort Monmouth, NJ 07703 RSARTORE-at-MONMOUTH-ETDL1.ARMY.MIL
Dear Tamara Howard, I'm not sure if this is exactly what you are looking for but I have seen people in Jeremy Pickett-Heaps' lab at Melb. Uni. use teflon powder to coat glass slides before embedding between them. They don't actually grow the cells on the slides but just use them for thin layer embedding of algae in Spurr's. I'm not sure where teflon powder comes from - sorry. Can you grow the cells on plastic coverslips that you can section? I too have experienced having coverslips smashing instead of peeling nicely from the resin and actually found patience, practice and the gentle persuasion of a single-edged razor blade angled down between the glass and the resin to yield enough glass-free resin + cells to make it worth the trouble. David Orlovich.
Tamara, Try growing your cells on Permanox plastic dishes. The plastic is resistant to ethanol, acetone and resin and the polymerized resin is easily separated from the petri dish as long as the resin isn't too thick. I usually use a resin layer of about half a mm. If you need to use glass coverslips, you can try pushing the still-warm coverslip against a block of dry ice. The differential contraction rates will sometimes free the resin but it isn't 100% effective.
Rod Kuehn University of Minnesota
On Wed, 19 Jan 1994, Tamara Howard wrote:
} Does anyone have any experience with cell monolayers grown on glass coverslips? } I've seen a method reported where you coat the coverslip with carbon before } adding the cells; this is supposed to allow the resin to be stripped from the } glass for sectioning. Does it work? We thought the regular culture } substrates would allow release, but the glass just breaks when we try to "peel" } the resin away. Any suggestions would be VERY HELPFUL...I'm trying to section } glass, now. } Thanks! } Tamara Howard } Pitt Med/Pathology } Pittsburgh, Pa } tah-at-med.pitt.edu
} One other question that I would like to direct to them or other } interested parties would be the requirement for installing a } "scrubber" on the exhaust fumes from the RIE. } ...The requirement for a "scrubber" on the exhaust has been } mentioned as a possibile environmental requirement. I have spoken } to some manufactures and they don't use scubbers due to low flow } rates in their machines and small quantities of gas involved. } } Thanks again. } } Richard Sartore } US Army Research LAboratory } AMSRL-EP-RA } Fort Monmouth, NJ 07703 } RSARTORE-at-MONMOUTH-ETDL1.ARMY.MIL
The requirement for a scubber is more likely to be a matter of law and agency regs (military regs in your case) than real need, BUT it may also depend on what's coming off of the targets. The gas would be no trouble (so the manufacturers aren't going to worry about it), but if you're etching something like Gallium Arsenide semiconductors, there'll be reactive arsenic ions and the like coming out; that could be a problem. Scrubbing should be easily and cheaply done be bubbling the exhaust through distilled water (with maybe cotton batting in the outlet to make sure), which would be then disposed of by the usual regs for chemical waste. Phil Oshel po-at-parmly1.parmly.luc.edu
Re} TEM biol. cells on glass Tamara, I assume that you must embed the cells as a monolayer so that you can either orientate them or locate specific cells. If so, then you can take any of the blocks that you have already embedded, place them in liquid nitrogen and then warm them up. Repeated cycles of cooling and warming will crack the glass and will often cause it to shoot off the plastic. You must remove all resin from the top side of the block beforehand and be careful that the glass pieces do not get into your eyes. We use this method routinely to locate, and section, single cells that have been microinjected. The cells are grown on a locator slide and the pattern is transferred to the resin. You can help the glass removal by scoring the coverslip with a diamond before cooling and do not worry if the resin breaks. You will usually be able to find what you want amongst the pieces.
Cutting plastic coverslips is not easy, but a viable alternative to consider, if you only want orientation, is to grow the cells on the special filters produced by Costar and Falcon. These embed well and can be sectioned in resin. They are more difficult to cut cryosection from, but even this is possible.
If you only want a pellet of cells then it is better to grow them in plastic dishes. You can remove them, before fixation, by treating them with proteinase K, and after fixation by scraping with either a soft wooden or teflon scraper (not the normal cell scrapers that are available).
Good Luck
Paul Webster Yale School of Medicine (203) 785 5072.
Tamara Howard asks about stripping processed cells off glass coverslips, and suggests using carbon coated coverslips.
I have 3 suggestions:
1. I seem to recall seeing a paper/idea/suggestion once somewhere (???...) that rapid temperature changes at the glass resin interface will help break off the coverslip cleanly. Place a solid (pre-filled with resin and then polymerized) gelatin/Beem capsule over the cells on the coverslip and then polymerize them, attaching the cells to the capsule. Later, rapidly cool the coverslip, twisting the beem capsule. Play around, and let me know what works ! *** ONE ADVANTAGE: You could also first polymerize the cells between 2 coverslips (the one that it was grown on, and the other greased with vaseline so that it can easily be disloged after polym.), so that you could observe the cells clearly under phase contrast, mark the cells of interest, and finally place a solid resin capsule over the marked area, thus selecting the cells of interest.
2. Use propylene oxide (PO). This works like a dream when processing cells in plastic culture dishes: do all your processing for TEM right up to full dehydration with ethanol in the dishes. Monolayers process (fixation and dehydration) very quickly, and so it is a real easy technique to try. Decant the 100% ethanol from the dish, then quickly pour on pure PO, gently tilt the dish once or twice, and as the plastic of the dish starts to dissolve, the whole monolayer "peels off" and can be decanted into a suitable tube or small vial. Replace PO with the first change of resin, and carry on, embedding the mat of cells, perhaps using gentle centrifugation for the pure resin changes. Dave Sanan, now somewhere in California (give me a call, man!) first showed me this trick.
Perhaps the PO could even lift monolayers off the glass ? Would certainly work for plastic coverslips.
3. Hydrofluoric acid. Here's the real McCoy! Ref: J. Microsc. 104: 205- 207. Use hydrofluoric acid to etch away the coverslips, leaving the resin and cells behind. Be real careful as HFA is very corrosive. Work in a polyethylene or polytetrafluoroethylene dish in a fumehood.
Good luck
Ian
*********** ************ Dr Ian S Harper Int. Tel #: 27-21 938 0347 Experimental Biology Programme Int. Fax #: 27-21 938 0456 Medical Research Council Internet: iharper-at-eagle.mrc.ac.za PO Box 19070 Tygerberg, 7505 South Africa *****************************************
Thanks to B.Miner for forwarding the useful comments from P.Catinella and B.Edwards on RIE etching microelectronic devices.
One other question that I would like to direct to them or other interested parties would be the requirement for installing a "scrubber" on the exhaust fumes from the RIE. We are working up a estimate for the possible total cost of purchasing/operating/maintaining a plasma etcher in our laborato- ry. The requirement for a "scrubber" on the exhaust has been mentioned as a possibile environmental requirement. I have spoken to some manufactures and they don't use scubbers due to low flow rates in their machines and small quantities of gas involved.
Thanks again.
Richard Sartore US Army Research LAboratory AMSRL-EP-RA Fort Monmouth, NJ 07703 RSARTORE-at-MONMOUTH-ETDL1.ARMY.MIL
---------------------------------------------------------------------------- ---------------------------------------------------------------------------- --------------------------------------- Richard, If you are only dealing with a low volume of gas and you have a fume hood near by then you can do as we do exhaust fumes through there as most fume hoods, I would imagine, have a scrubber attached.
we only use plasma etching techniques for "deprossessing" integrated circuits for failure analysis so our gas flow is minimal.
For embedding monolayers we have been growing cells on a polymer film in place of glass cover slips. It is called Aclar and is available (in large quantities only) from ProPlastics, Linden NJ. It is optically very clear and separates easily from all embedding resins. It is apparently a Teflon- like substance. I was required to purchase more than a lifetime supply, so anyone who would like to try a sample should contact me. Not all cell lines will grow on the naked stuff. Some require a collagen coat and others never grow at all as they do on polystyrene dishes. or glass.
FROM THE PRESIDENT OF THE ROYAL MICROSCOPICAL SOCIETY
Something for Nothing! Bursaries Bursaries Bursaries ************************************************************************ Over many years it has never ceased to amaze me that RMS bursaries, to help eligible people attend meetings or courses, are not snapped up. It has even been known for them not to be fully taken up by the date of the sponsored event. Although admittedly the value of the bursary may not cover all of the expenses incurred in attending an event, they can be very useful primers in persuading other sponsors to make a contribution. Anyway, we announce here, bursaries for events in 1994.
RMS International Bursaries in Microscopy ************************************************************************ The RMS International Bursaries are intended to help young microscopists working outside Western Europe to attend RMS Courses or Conferences. The awards will be made to help with the registration and accommodation costs but it will not normally be possible to help with the cost of travel to the United Kingdom. The Society will offer up to six Bursaries annually and it is unlikely that any single award will exceed œ250. There are no strict rules or definite age limits but it is likely that they will be made to assist young scientists who would otherwise be unable to attend the Course or Conference. The Bursaries are not limited to Fellows or Student Members of the RMS, but it is unlikely that an award will be made to an Applicant currently working in North America or Japan.
The application for an RMS bursary can be made at any time, but should be made as far in advance of the Course or Conference as possible. The application should include details of the Course or Conference to be attended, a copy of any abstract(s) to be submitted and also a copy of the Applicant's Curriculum Vitae and publication list (if appropriate). The application should be accompanied by a letter of support from the Applicant's Head of Department or from a Fellow of the Royal Microscopical Society.
It is expected that the Applicant will have made efforts to find funding from elsewhere and he/she will be expected to show that such an application has been made - even if it was not successful.
RMS Bursaries in Microscopy ************************************************************************ The RMS bursaries will only be available to Fellows or Student Members of the Society. Non-members of the RMS are not eligible. The awards will be made to help with the registration costs of a Course or Conference, but it will not normally be possible to help with the cost of travel or accommodation. There are no definite age limits, but it is likely that they will be made to assist young microscopists who would otherwise be unable to attend. It is unlikely that any single award will exceed œ250 nor be made to an Applicant currently working in North America or Japan.
The application for an RMS bursary can be made at any time, but should be made as far in advance of the Course or Conference as possible as funding may be limited and to allow for fair refereeing of the applications.
It is expected that the Applicant will have made efforts to find funding from elsewhere and he/she will be expected to show that such an application has been made - even if it was not successful.
Applicants are advised that preferential consideration will be given to DipRMS and TechRMS candidates and those who have no financial support from their employer or other source. Please note that each application must be endorsed by the Applicant's Head of Department or employer.
The names of the successful Applicants will be published in the RMS Proceedings.
Application Forms for RMS Bursaries are available from: The Administrator, Royal Microscopical Society, 37/38 St Clements, Oxford OX4 1AJ, United Kingdom. Completed applications must be returned to the Administrator at the same address.
The Summer School in Light Microscopy for 1994 will be organised on a modular basis, as in 1993, to allow flexibility in matching the needs and interests of participants to the subject matter provided, and to make it possible to include minor or specialist topics for which the demand might not justify the provision of separate courses. These topics can be offered in conjunction with others and thereby share the availability of instructors and expensive up-to-date equipment. There will be evening lectures and discussions to obtain the best use of the time available, and the numbers of participants will be strictly limited.
The first three days (Sunday evening to Wednesday evening), the Principles of Light Microscopy module will consist of lectures, demonstrations and practical classes on the fundamental aspects of light microscopy and the various imaging and contrast modes which can be used in the observation and characterisation of biological specimens, polymers, ceramics, minerals and metals. This module will cover the basic concepts of all forms of microscopy, leading on to the functioning and limitations of the light microscope, and introducing techniques for enhancing contrast. Practical work will make use of a variety of types of microscope from the major manufacturers, and the module will provide a practical understanding of the phenomena which lie behind the principles and practice of light microscopy.
From Thursday morning, the course will be divided into three specialist modules from which participants may select. Analytical and Applied Microscopy is an extension of the Principles module, these two together covering approximately the same ground as our former one-week Principles course. It will build on the topics covered in the Principles module and will show how they may be applied in practice. This will assist the development of a systematic and analytical approach to microscopy. A workshop format will enable participants to examine and discuss the images obtained from samples which will be provided and their own specimens, using a wide variety of techniques. The emphasis will be on the correct adjustment of the instruments, the strengths and weaknesses of each technique and the interpretation of images. This module will be especially valuable for microscopists working in industrial laboratories.
The Polarised Light module will address itself to the interpretation of contrast, not only in ceramics and minerals, but also in biological materials in which components of the structure are birefringent. It will attempt to explain by the use of simple diagrams and demonstrations, and with the minimum use of mathematics, the colour changes which are observed, and how contrast may be enhanced by the use of accessories. This module is designed to be of use to biologists, materials scientists and geologists.
In the Image Recording module it will be assumed that participants have a thorough understanding of the techniques of imaging and contrast enhancement. The module will include the principles of photography and video imaging, the transfer of the image from the microscope to the camera, and the design and construction of image-recording equipment. Time will be available for exposing black-and-white and colour film, which will be processed and evaluated before the end of the module.
Participants may register for the whole week (for the Principles module and one of the three specialist modules), for the Principles module alone, or for one of the specialised modules alone. Because the specialised modules are designed to build upon the groundwork presented in the Principles module, we expect that participants in specialised modules will normally either attend the Principles module at the beginning of the week, or have attended a previous RMS Light Microscopy Summer School.
We anticipate the customary extremely generous provision of equipment and materials from the manufacturers, for all parts of the course.
Principles of Light Microscopy Sunday evening 17 July to Wednesday evening 20 July 1994 (a 3 day module)
This module will consist of lectures, demonstrations and practical work on:
The history of microscopy. Introduction to microscopy. Limitations of the eye. Resolution, Contrast, Magnification. Interactions between light and matter. Refraction and lenses, geometrical optics, conjugate planes. Aperture. Illumination of the specimen in transmitted and reflected light.
Lens aberrations and their correction; the choice of optical components. Diffraction and its consequences for the microscope image. Generation of contrast. Introduction to bright-field, dark ground, phase contrast, polarized light, differential interference contrast and fluorescence.
This module will adopt a workshop approach and consist of short informal lectures, demonstrations and practical sessions. It will cover the correct adjustment of the microscope, the strengths and weaknesses of each technique, and the interpretation of images, in both transmitted and reflected light. Participants will have the opportunity to become familiar with techniques of their own choice according to their interests, using where possible, their own specimens. Facilities for specimen preparation will, however, be limited. We expect equipment for the following techniques to be available:
This module is designed to introduce biologists, materials scientists and geologists to the usefulness of polarized light techniques, and will involve the minimum use of mathematics. Lectures, demonstrations and practical work will cover the following topics:
The nature of light and polarization. The interaction of birefringent materials with plane-polarized light. Stress optical effects, the use of accessories and compensating plates. Minerals, ceramics, biological materials and fibres. Reflected polarized light techniques. Introduction to conoscopic techniques.
This module will discuss the recording of images by drawing, by photography and by video techniques. Equipment and materials will be available to enable participants to record images in black and white and colour of their own specimens and to discuss difficulties and results. The following topics will be covered:
Drawing, photography and video methods; an overview. Transferring the microscope image to the film and video camera. Monochrome and colour film; basic principles, use and processing. Negative and reversal films. Printing. 'Instant' monochrome and colour films. Colour temperature, light sources, films and filters. Equipment for photomicrography; focusing and determining exposure. Photomacrography. Video cameras, monitors and printers.
TO OBTAIN FURTHER DETAILS OF THIS COURSE SEND AN EMAIL MESSAGE WITH YOUR NAME AND ADDRESS TO RMS-at-UK.AC.OX.VAX, OR CONTACT THE RMS ON TELEPHONE +44 (0)865 2488768, FAX +44 (0)865 791237.
A range of techniques have been devised for localising antigens at the electron microscope level. One of the difficulties commonly faced is the choice of which method to use with any particular antigen-antibody combination. The aim of the course is to provide a theoretical and practical introduction to the various methods available and covers antigen location, using both transmission and scanning electron microscopy.
The advantages and disadvantages of the various methods available will be discussed, but the emphasis will be on the practical aspects of the techniques and will provide experience of the following methods: pre-embedding labelling for scanning electron microscopy; low temperature embedding in Lowicryl resins; preparation and labelling of both resin sections and thawed cyrosections; rapid freezing by impact and high pressure followed by freeze substitution and low temperature embedding; silver enhancement of colloidal gold for light and electron microscopy; immunolabelling of 1æm resin sections; and epi- polarised light microscopy. Registrants are encouraged to discuss specific problems with the course organisers and if possible, to bring samples with them for processing and labelling during the practical sessions, which will be supported by Leica (UK) Ltd.
The course is primarily aimed at electron microscopists with experience of routine processing methodology who wish to become familiar with ultrastructural immunocytochemical labelling techniques.
The practical nature of the course, means that numbers will be restricted to a maximum of 10 registrants.
FOR FURTHER DETAILS CONTACT THE RMS - TELEPHONE +44 (0)865 248768, FAX +44 (0)865 791237, EMAIL RMS-at-UK.AC.OX.VAX.
The unprecedented upsurge in the application of immunocytochemistry in the life sciences during the past decade continues with increasing vigour. Both light and electron microscopy are important techniques in routine diagnosis and research in medicine and biology. This technology has contributed so much towards our current understanding of the cell, that the modern microscopist must be part immunologist as well as being skilled in microscopy.
The underlying principles of immunocytochemistry apply equally to light and electron microscopy. This five-day course has been specially designed to utilise this overlap and therefore, will be of value to both light and electron microscopists. The course is structured towards a technical appreciation of immunocytochemical techniques, since once they are mastered, they can be applied to any system. Each year this popular course is updated in the light of new developments in immunocytochemistry and is also suitable for participants interested in the plant sciences, as we teach some of the specialist techniques required for the handling of plant cells. The course will be of immense value to any life science microscopist/cell biologist of any background, who intends to use or is starting to use immunocytochemistry for routine purposes or research.
The course counts towards qualification for the Diploma of the Royal Microscopical Society.
The main emphasis of the week will be to give participants sufficient practical experience and knowledge of immunocytochemistry to carry out immunolabelling in their own laboratories. At the same time, a series of lectures will be given by more specialist exponents in various areas of immunocytochemistry.
The three practical days will be led by experts in their particular field: Tony Leathem and Susan Brookes (LM immunocytochemistry), Paul Monaghan (immunogold labelling for EM) and David Hughes (silver enhancement). After an introduction on the biology and production of antibodies, the practicals will be backed up with lectures on the applied aspects of the respective techniques. Of particular interest is the series of specialist lectures which will include, botanical immunocytochemistry (Chris Hawes), in situ hybridisation (John Davies), confocal microscopy (Mark Fricker), immuno-scanning EM and special applications of colloidal gold. An evening workshop will be held to introduce cryo-techniques in immunocytochemistry and to demonstrate some of the latest equipment used in low temperature tissue preparation.
FOR FURTHER DETAILS CONTACT THE RMS - TELEPHONE +44 (0)865 248768, FAX +44 (0)865 791237, EMAIL RMS-at-UK.AC.OX.VAX.
Last year we experimented with a new format for our Flow Cytometry course. Following its success both in attracting students and instructing and informing them, we will continue with the new format. There will be two courses - basic and advanced - run sequentially to give the potential student the choice of attending either or both of the courses. We anticipate that there will be at least three bench top cytometers for use by the students and one machine equipped with two lasers for the advanced course. As usual, we will rely on the generous co- operation of the manufacturers, Becton-Dickinson, Coulter and Ortho, who lend us both the machines and experienced operators to run them.
The basic course will assume little prior experience and take the student through the most important applications. Although it will fill the needs of someone working in a research environment, it will have a slight clinical bias. The majority of bench top flow cytometers are now to be found in clinical laboratories.
The first day will consist of a series of talks describing the basics of flow cytometry. A simple practical (measurement of a DNA histogram from cultured cells) will serve as an introduction to using the bench top flow cytometers.
There will be two practicals on the second day. In the first, students will use antibodies labelled with three different fluorochromes to identify lymphocytes subsets in human peripheral blood. This practical will also demonstrate the importance of using light scatter to separate lymphocytes, monocytes and granulocytes in samples of blood. In the second practical (lead by Richard Camplejohn from St. Thomas's Hospital), nuclei will be extracted from a formalin-fixed, paraffin embedded tumour and the DNA histogram recorded. The rest of the programme will include lectures on the analysis of DNA from clinical samples and on immunophenotyping in a hospital laboratory.
The third and final day will also be the first day of the advanced course. In the practical demonstration, lead by George Wilson (GRC Gray Laboratories), students will investigate the cell cycle kinetics of a mouse tumour which will have been labelled in vivo with a thymidine analogue (bromodeoxyuridine, BrdU). There will be a lecture on applying this technique and its clinical application and on DNA measurement and cell cycle analysis, the principles of cell sorting and the measurement of antigens associated with cell proliferation.
The advanced course will join us for the last day of the basic course (see above). On their second day, they will prepare and analyse chromosomes, using both bivariate and univariate analysis. We will set up a flow cytometer in the lecture theatre and project the computer screen so that it can be seen by the whole class. Jim Watson (MRC, Addenbrookes, Cambridge) will lecture on time as a parameter and then run experiments in the lecture theatre on intracellular enzyme kinetics looking at esterases and glutathione-S-transferase. A lecture on the measurement of intracellular pH and calcium ions will also be followed by a live demonstration. The other lectures will be on chromosome analysis and sorting and on further applications in cell and molecular biology.
The third and final day will consist of lectures on studying apoptosis, measuring multi-drug resistance in tumours and lectures and practical demonstrations on the measurement of cell cycle kinetics using the BrdU-Hoechst/PI method and on the interaction of fluorochromes and DNA.
We expect that anyone attending our basic course will come away with a sound grasp of the principles of flow cytometry, an understanding of the concepts behind data analysis, a feel for some of the problems and the confidence to run the commoner applications. Everything in the course will be relevant to workers in a clinical environment.
The advanced course will be at the forefront of the technology. It should give students a broad understanding of the wide range of applications of flow cytometry in a modern research laboratory. It will help them to establish new techniques and ought to give them ideas for new experiments in their own laboratories .
We hope that many students will take the opportunity to stay the whole week and to benefit from a broad look at flow cytometry - from basics to the most advanced applications. The RMS course is the only comprehensive course on flow cytometry to be run in the UK
FOR FURTHER DETAILS CONTACT THE RMS - TELEPHONE +44 (0)865 248768, FAX +44 (0)865 791237, EMAIL RMS-at-UK.AC.OX.VAX
We have versions of Diffract, DiffractII, and Desktop Microscopist. The best one is the Desktop Microscopist by far. It works much better, doesn't have as many faults as diffract and is more user friendly. It doesn't have that annoying propensity to crash all the time as diffract.
I would like to use freeze substitution for a low magnification TEM study of cell-extracellular matrix relationships in amphibian embryos. Can anyone suggest an appropriate fixative and concentration for adding to the sustitution medium (MeOH) prior to embedding in EPON?
Dave Parichy Section of Evolution and Ecology Univ. of California, Davis 916 752 3634
Mime-Version: 1.0 Content-Type: text/plain; charset="us-ascii" To: "Rick A. Harris" {szrick-at-bullwinkle.ucdavis.edu}
} I would like to use freeze substitution for a low magnification TEM study } of cell-extracellular matrix relationships in amphibian embryos. Can } anyone suggest an appropriate fixative and concentration for adding to } the sustitution medium (MeOH) prior to embedding in EPON? } } Dave Parichy } Section of Evolution and Ecology } Univ. of California, Davis } 916 752 3634
You might find useful information in:
Allanspach, A. 1993. Ultrastructure of early chick embryos after high pressure freezing and freeze substitution. Micoscr. Res. Techn. 24:369-384.
Hippe-Sanwald, S. 1993. Impact of freeze substitution on bioolgical electron microscopy. Microsc. Res. Techn. 24:400-422.
Hunziker, E.B. 1993. Application of cryotechniques in cartilage tissue preservation and immunoelectron microscopy: potentials and problems. Microsc. Res. Techn. 24:457-464.
McDonald, K. and M. Morphew. 1993. Improved preservation of ultrastructure in difficult-to-fix organisms by high pressure freezing and freeze substitution: I. Drosophila melonagaster and Strongylocentrotus purpuratus embryos. Microsc. Res. Techn. 24:465-473.
I have just had a look in my favourite freeze-substitution book (Cryotechniques in Biological Electron Microscopy, eds: R A Steinbrecht and K Zierold) and they suggest the following freeze-substitution medium for methanol substitution:
Methanol containing 0.5% uranyl acetate, 1% OsO4, 3% glutaraldehyde and 3% water (the water comes from the 50% glutaraldehyde they used).
It actually looks a bit complex to make the solution up - they suggest:
add 9 mL of 50% aq. glutaraldehyde (in a liquid nitrogen precooled flask) to 60 mL methanol, then 3 mL of a 20% (w/v) soln of uranyl acetate in methanol are added; in a second precooled flask 1.5 g osmium tetroxide are dissolved in 75 mL methanol; both flasks are cooled to about 220 K, their contents poured together and vigorously shaken.
They say that the mixture is highly reactive even at 240 K and so should be used in a few hours.
Substitution time was 8 hours each at -95 deg C, -60 deg C and -30 deg C. Then 30 minutes at 0 deg C. Replace the substitution mix with pure acetone (still at 0 deg C), warm to 7 deg C and infiltrate with araldite/Epon.
The original reference to this protocol is: Muller, M., Marti, T., and Kriz, S. (1980). Improved structural preservation by freeze-substitution. In: Brederoo, P., and de Priester, W. (eds). Electron Microscopy 1980, vol.II. Proc. 7th Eur. Congr. Electron Microsc., Leiden, pp. 720-721.
I've always used 2% OsO4 in acetone at -70 deg C for up to 7 days and then embedded in Spurr's resin. The best reference I have for that is:
Howard RJ and O'Donnell KL (1987). Freeze-substitution of fungi for cytological analysis. Exp. Mycol. 11:250-269.
I hope this is of some use.
David Orlovich School of Biological Science University of NSW PO Box 1 Kensington NSW 2033 Australia
Subject: Time:2:18 PM OFFICE MEMO RE} SEM RES STD Date:1/24/94 The method I have used with good success was published on p. 68 of the May 1987 issue of the EMSA Bulletin. If you don't have access to that issue of the Bulletin, send me your FAX address and I'll send a copy to you. Bigelow-at-umich.edu.
I am looking for software (vendor / public domain) that can draw crystal structures and print these diagrams on a laser printer. I would like to be able to enter the crystal system, cell edges, etc.. and have the diagram show the coordination of differing cation sites. Any suggestions?
Message-Id: {MAILQUEUE-101.940125090324.512-at-FS-IAM-1.JRC.NL} To: Microscopy-at-anlemc.msd.anl.gov
I intend to do some internal strain measurements using HOLTZ lines, so could anyone suggest any software which allows the simulation of HOLTZ lines, and where I could find it?
Kodak has discontinued direct duplicating film 2468 in 100 foot rolls. We have used this film successfully for years to make presentation slides from halftone prints. What should we use now as a replacement?
Received: from MAILQUEUE by MICROBIO1 (Mercury 1.11); Tue, 25 Jan 94 11:24:30 EST Return-path: {EMLAB-at-opus.mco.edu} Received: from emoryu1.cc.emory.edu by transporter.microbio.emory.edu (Mercury 1.11); Tue, 25 Jan 94 11:24:27 EST Received: from anlemc.msd.anl.gov by emoryu1.cc.emory.edu (5.65/Emory_cc.3.4.12) via SMTP id AA15050 ; Tue, 25 Jan 94 10:51:32 -0500 Return-Path: EMLAB-at-opus.mco.edu
Kodak has discontinued direct duplicating film 2468 in 100 foot rolls. We have used this film successfully for years to make presentation slides from halftone prints. What should we use now as a replacement?
KODAK OFFERS A FILM CALLED " RAPID PROCESS COPY " (CAT. NO 174 6031 FOR THE 150 ft ROLL) WHICH IS A DIRECT REVERSAL FILM DESIGNED FOR THE PURPOSE YOU DISCRIBE. THIS FILM IS QUITE SLOW : EXPOSURES IN THE 30 TO 45 SECOND RANGE AT f 4.0 ON MY SETUP. ONE NEEDS TO CALIBRATE THEIR COPY STAND SETUP TO THE FILM AND PROCESSING.
THE PROCESSING IS VERY SIMPLE: DK 50 DEVLOPER FOR 10 MINUTES, FOLLOWED BY STOP AND FIXER.
I HAVE BEEN USING THIS FILM SINCE THE MID- EIGHTIES WITH EXTREMELY GOOD RESULTS. ANY QUESTIONS? CALL ME. lmelsen-at-unix.cc.emory.edu
Via: uk.ac.birmingham.computer-centre.ibm3090; Tue, 25 Jan 1994 16:27:15 +0000
On Tue, 25 Jan 1994 MACLAREI-at-ibm3090.computer-centre.birmingham.ac.uk wrote:
} I wonder if anyone has any good ideas on how to electropolish commercial } purity Titanium without getting precipitation of hydride needles. } } I have tried using a variety of acid based solutions, most of which } contained perchloric acid in varying concentration. All of these, however, } result in hydride precipitation. } } The net result of all of this is that I find it very difficult to prepare } decent specimens of Titatium by electropolishing. If anyone has any good } ideas as to how these problems can be overcome I would be very glad to hear } from you. } } Ian MacLaren
Ian,
You may want to try the following, (although I have NOT tried this particular recipe):
Ethanol (96%)................90 ml n-Butyl alcohol..............10 ml Aluminum Chloride.............6 g Zinc Chloride................28 g for 1-6 minutes; 20 -25 V dc; stainless steel cathode; room temp.; need to keep agitated (the solution, NOT the user {grin!} ) to prevent a passivating layer from forming: can use a stirrer, or oscillate the anode quite rapidly at a fixed distance from the cathode (approx. 1 to 2 cm). * remember: nicely TOXIC!!! {ref.: The Electrolytic and Chemical Polishing of Metals; Tegart; 1959; Pergamon Press}
Hope this is helpful, Rob
X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X X Rob Tayloe X MSM Spelunkers Club X X Metallographic Lab. X Missouri Speleological Survey /-v-\ \-v-/ X X Rolla Research Center X Bat Conservation International X X U.S. Bureau of Mines X Missouri Cave & Karst Conservancy \-v-/ X X tayloe-at-rorc.usbm.gov X National Speleological Society #32993 /-v-\ X X (314) 364-3169 x247 X American Cave Conservation Association X X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X\X
I think there is new chemistry for using T MAX films for direct positives. Someone from Kodak should pick up on this message and let the rest of us know about it. ***************************** * Greg Erdos * * Director, ICBR EMCL * * University of Florida * * Gainesville, FL 32611 * * gwerdos-at-gnv.ifas.ufl.edu * * 904-392-1295 * *****************************
I called my supplier of 2468 today. They say that the film has NOT been discontinued. (And this company claims they are the only E. Coast supplier!) What's the truth???
If your prints come from large-format negatives, you can photograph the negatives on a light box with technical pan or t-max. If you need a direct positive film, you can use Ektachrome color slides or use t-max with a direct-positive developing kit.
Rod Kuehn University of Minnestota
On Tue, 25 Jan 1994 EMLAB-at-opus.mco.edu wrote:
} Kodak has discontinued direct duplicating film 2468 in 100 foot rolls. } We have used this film successfully for years to make presentation } slides from halftone prints. What should we use now as a replacement? }
Return-path: {BERGRH-at-MSUVX1.MEMST.EDU} Received: from memstvx1.memst.edu by gnv.ifas.ufl.edu (PMDF V4.2-11 #3240) id {01H848IIDF408WW4DH-at-gnv.ifas.ufl.edu} ; Tue, 25 Jan 1994 23:26:34 EST Received: from MSUVX1.MEMST.EDU by MSUVX1.MEMST.EDU (PMDF V4.2-14 #5958) id {01H845ET8EPC9BWPOF-at-MSUVX1.MEMST.EDU} ; Tue, 25 Jan 1994 22:28:15 CST
Greg-- I am reading your Microscopy post and, not adept at replying to these public forums, am writing you direct. I used the TMax direct reversal kit for my talk at this summer's MSA meeting and was very pleased with the quality of the slides it produced. I made "superslides" by using (hard to find) 127 film mounts and TMax 100 film size 120. The kit specifies rating the film at ASA 50, half its normal speed. The kit instructions are well written and I got good results from the first roll on. Instructions for extending development with subsequent rolls are clearly written--as I recall the kit will process about 8 or 10 rolls and cost me $35 (?). Significantly more rolls of 35mm would be possible
Direct Pos. Film users: Kodak makes a TMax direct postive film developing kit for making black and white slides. Catalog number 812 1188. Cost: approx. $30. When doing line copy I have also used LPD4 film for continuos tone copy and have had very good results. There's a little trickery involved but it allows you to do line and continuous copy on the same roll of film. If anyone is interested, I'll tell how it is done. Phil Rutledge prutle1-at-gl.umbc.edu
Message-Id: {MAILQUEUE-101.940126090559.480-at-parmly1.parmly.luc.edu} To: microscopy-at-anlemc.msd.anl.gov
} Date: Wed, 26 Jan 1994 09:26:33 -0500 (EST) } From: rutledge phil {prutle1-at-gl.umbc.edu} } Subject: Tmax } To: microscopy-at-anlemc.msd.anl.gov
} Direct Pos. Film users: } Kodak makes a TMax direct postive film developing kit for making black } and white slides. Catalog number 812 1188. Cost: approx. $30. When doing } line copy I have also used LPD4 film for continuos tone copy and have had } very good results. There's a little trickery involved but it allows you } to do line and continuous copy on the same roll of film. If anyone is } interested, I'll tell how it is done. } Phil Rutledge } prutle1-at-gl.umbc.edu
I'll second this--I've used LPD-4 for making direct-positives of photographs for slides when I only want to mess with developing one type of film. 8 secs. at 1/2-stop intervals from f111/2 to f4 or 41/2 will usually get you a usable frame. This range of f-stops is conservative; actually shouldn't need more than a couple of brackets once you figure out your local conditions. The Direct Positive sold by Ted Pella does do a better job, but.... Phil Oshel
Kodak make a T-Max Reversal Kit for direct positives. The catalogue number is K-8121188. It comes as a 1 quart unit. In Canada it sells for about $35.00.
Kodak make a T-Max Reversal Kit for direct positives. The catalogue number is K-8121188. It comes as a 1 quart unit. In Canada it sells for about $35.00.
} } I have found something that I think works better, but takes a day to } develop - regular color Ectachrome slide film (Tungsten). The results } are very good and having somebody else fool with the wet chemistry is } much better. I think that the development process is C-47 which is the } same for color negatives which means that you can have it done at 1 hour } photo shops. As a reult, I only use the MP 5360 when I have an } emergency. } } } Scott Walck } walcksd-at-ml.wpafb.af.mil } Materials Directorate } Wright Patterson AFB, OH
I have had excellent results with Ektachrome, but find it takes 25-30 minutes to development: process E-6 with the Kodak hobbyist pack, 10-30 minutes to dry, depending on if you have a film drier. Most 1-hr photo-shops that I know of won't do E-6 films. Phil Oshel po-at-parmly1.parmly.luc.edu Parmly Hearing Inst. see c. v. on MSA Bulletin Board. Please!
Slides I can't believe that anyone is satisfied with the 35mm format for projection slides. Besides, copying prints with a camera always results in a loss of quality. The best way is to take the information straight from the negative, although this does not let you label the image with letraset. I print directly onto film, using an enlarger and mount the image in the super-slide frames (40x40mm) loved by electron microscopists and hated by projectionists. The image can be carefully framed and the contrast can be easily manipulated. The film to use is Agfa sheet film, so you will have to find your own supplier, and can be purchased in boxes of either 10x8 or 10x12. Use it as you would paper under an enlarger. For a soft image use "Litex premium camera film 0910P". For a harder finish use "Litex camera halftone film 0811P". These films are normally developed in Gevaline G7C developer, also from Agfa, which will produce high contrast but they can also be developed in D-19 for a softer contrast. I am sure that acutance developers will also work well. The film can only be handled in red light and has to be dish developed but the results are worth it. Try it and compare with camera copies. There is nothing like a big image to make your point. If you are interested and require more details then feel free to contact me. Paul Webster Yale School of Medicine paul_webster-at-quickmail.yale.edu (203) 785 5072.
} Slides } I can't believe that anyone is satisfied with the 35mm format for projection } slides. Price! Money! Grant funds! 35mm is cheap! And everyone has a slide projector that will take 35mm. Phil Oshel
I would appreciate any feedback from users of Digital confocal software packages like MICRTOME from VayTek or comparable systems.
1) Does cost justify capabilities.
2) How it compares to laser systems?
3) Are manus userfriendly?
4) If you used package already would you buy an upgrade or move on to a different system?
Thanks.
Cesar D. Fermin, Ph.D. Tulane University Medical School Department of Pathology 1430 Tulane Ave/SL79 New Orleans, La 70112-2699 (504) 584-2521 Fax 587-7389
"Price! Money! Grant funds! 35mm is cheap! And everyone has a slide projector that will take 35mm. Phil Oshel"
What a rude reply Phil! I don't deserve that.
Most EM labs have a darkroom with an enlarger. A box of 100 sheets of film is cheap and will last for years (I bought my boxes four years ago and still have enough to send to you, Phil Oshel, if you want to try out the system. The 40x40mm slide holders (from GePe) fit into a 35 mm slide projector -they just use up more of the available space. If anything it is all cheaper than buying a copystand and 35 mm camera. Why do I bother?
During my short stay in Sendai, Japan, I learned a good way to make overheads from electron microscope negatives. The material is FUJIGRAPH PROJECTION FILM PT-100, which comes at least in size 21x29,7cm (DIN A4) in 100 sheets packages. This film can be used just like the enlarging paper in the darkroom. The results are far better than with any oldfashioned way. These overheads are good for teaching purposes because you can easily make markings on them.
Regards, J M
- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - Jouko K. Maki Navigare necesse est... Laboratory Manager, Ph.D. Laboratory of Electron Microscopy Kiinamyllynkatu 10 FIN-20520 TURKU FINLAND University of Turku Tel.: + 358 21 633 7318 INTERNET: jouko.maki-at-utu.fi FAX.: + 358 21 633 7380
} Re} Slides } } "Price! Money! Grant funds! 35mm is cheap! } And everyone has a slide projector that will take 35mm. } Phil Oshel" } } What a rude reply Phil! I don't deserve that.
My apologies...but I repeatedly run into people, usually from medical schools, who don't understand why others don't use this or that, usually expensive, technology, instead of all the old stuff. The litany can get very frustrating. The answer is usually money. And often administrators. Labs and smally schools have photographic equipment & 35mm cameras, 35mm slide mounts, etc.. I have worked in EM labs for 13+ years, and have spent many hours all of the equipment you mention below. I have had much trouble with 40X40 slide holders jamming in 35mm projectors, but haven't tried the brand you recommend. Plus, multiple-image plates & 4"X5" Polaroid images still need to be photographed on the copy stand. And a good 35mm camera is still the cheapest, easiest way to produce the slides. Pros don't use them out of tradition. Personally, I'd prefer to use 120 film, or even 4X5, but....And most light microscopes come with 35mm camera backs. Meaning you need all that stuff anyway if you're a service lab, or anyway, not a dedicated lab where only one photpgraphic system is used (which most are). But what type of film comes in 100 sheet boxes & lasts for years? The film I've seen & used in TEMs & any other microscope--sheet, 35mm, or 120 roll--goes quickly. Printing paper's worse. Especially with students and EM courses! Thank you for the offer to send some materials, but it'd be a loss now--our grant & my job got cut, so it wouldn't be used. Phil Oshel } Most EM labs have a darkroom with an enlarger. A box of 100 sheets of film is } cheap and will last for years (I bought my boxes four years ago and still have } enough to send to you, Phil Oshel, if you want to try out the system. The } 40x40mm slide holders (from GePe) fit into a 35 mm slide projector -they just } use up more of the available space. If anything it is all cheaper than buying } a copystand and 35 mm camera. Why do I bother? } } }
To E'mers wanting EM slides: The easiest way to make EM slides is to put your neg in an enlarger and shoot onto another piece of EM film (type 4489). Develope it as you would normally process em film. This allows you to develope in a tray under a red safelight and allows control of the density of the slide. It gives great contrast in the slide or you can control the contrast by the way you develope the film. I started doing it this way for em slides only over 25 years ago and have always gotten good contrast whether the original negative was on the flat or contrasty side. Phil Rutledge prutle1-at-gl.umbc.edu
As I said in a previous message, I use LPD-4 for continuous tone slides on the same roll of film when I am doing line copy. The way I do this is as follows: I use a Polaroid MP-4 copy stand equipped with a Nikon F3 and a 90mm macro lens.
Line Copy __________
Exposure: 5 seconds/f:6 Develope: D-19 full strength- 2 minutes Fix: 2 minutes in rapid fix with Orbit Bath added. Orbit Bath allows a 2 minute fixing time and a 5 minute wash. It's better than Kodaks Hypo Clearing Agent. Wash: 5 minutes, dry, mount
Continuous Tone ________________
This requires a little playing with to determine your exposure. I use same exposure for the line copy but I add 3-5 seconds more. I pre-fog the film by exposing the copy for 8-10 seconds. At the same time I continuously move a grey scale card under the lens until I get to the 5 second mark for the line copy exposure. Process as above. I have always had good results with this method. You just need a little coordination exposing this way. Phil Rutledge prutle1-at-gl.umbc.edu
This is the text of an advertisement that will appear in the british press next week for a postdoctoral position which will become vacant on the 1st of June this year. We would like to identify an appropriate candidate as soon as possible - hence the short deadline. Applications will be accepted by FAX but NOT BY ELECTRONIC MAIL. Please note that, due to circumstances beyond our control, we will have to give preference to candidates who are citizens of nations in the European Economic Community.
The University of Birmingham School of Metallurgy and Materials
POSTDOCTORAL RESEARCH FELLOWSHIP
Applications are invited for the above post to work on a 2.5 -year SERC-funded project entitled "Mechanical Behaviour of Nb3Al Alloys". The successful applicant will investigate the basic defect structure and deformation mechanisms in these alloys and to explore strategies for enhancing their ductility.
A Ph.D. degree, extensive experience of electron microscopy techniques and familiarity with the physical metallurgy of intermetallic compounds are essential.
Preliminary enquiries should be directed to Dr. M. Aindow - Tel: (44) 21 414 5188, FAX: (44) 21 414 5232, Email M.AINDOW-at-BHAM.AC.UK, or Dr. I.P. Jones - Tel: (44) 21 414 5184.
Application forms and further particulars may be obtained from The Director, Staffing Services, The University of Birmingham, Edgbaston, Birmingham, B15 2TT, United Kingdom or telephone (44) 21 414 6483 (24 hours) and quote reference G10613/94. The closing date for receipt of applications is 18/2/94.
The University of Birmingham is an equal opportunity employer.
Mark Aindow, School of Metallurgy and Materials, Telephone; (021) 414 5188 The University of Birmingham, FAX; (021) 414 5232 Elms Road, Edgbaston, Birmingham, Email; M.AINDOW-at-BHAM.AC.UK GB B15 2TT, United Kingdom.
I have had quite a few requests asking how to access the new sci.techniques.microscopy newsgroup. So, I thought I would post a summary of the whats, wheres and hows of Usenet News, as I understand them, to the NIH-Image and Microscopy mailing list.
What is Usenet News? Usenet News, unlike email, is not distributed to an individuals account, but rather to a server at a particular institution (be it a government lab, university or private company) that can then be accessed by individuals at that institution. Usenet News is a system where messages are posted to newsgroups that are designed to cover a particular topic. Examples of topics are rec.windsurfing (recreational: windsurfing), comp.sys.mac.digest (Computer: Macintosh question and answer digest), sci.materials (Science: Materials Science) and, of course, sci.techniques.microscopy (Science: Microscopy Techniques of all kinds). There are many hundreds of worldwide newsgroups, i.e. groups that are distributed around the world, and then there also are groups that are only distributed locally. Examples of world-wide groups are those I have mentioned above and here at the University of Michigan we have local groups for class disscussion. As an example, for our electron microscopy course (MSE 562) we could have a group umich.eng.mse562 and it would only be available to readers in the umich.edu domain.
Postings to each newsgroup are stored on the institution's local NNTP (sorry guys I dont what this is an accronym for unless it is Network News Transport Protocol!) server and distributed to other sites. If the same kind of system is used now as was used when Usenet was first started, again I am no expert here - just a user, then each news server will exchange messages with a number of other servers geogrpahically close to it and the postings will sort of hop from one system to another and propgate around the world. When Usenet was first started the transfer was by dial-up modem lines, now the how system uses the Internet. If there is someone out there who can describe this process more exactly, please let me know or post a summary.
How do I read Usenet News and post to Newsgroups? If you want to use Usenet News you need access to an NNTP server and news reading/posting software to run on your local computer, workstation or terminal. You should contact your local network adminsitrator and ask them if you have access to a Usenet feed and which net-news software they recommend for the machine you use. If they do not have access, ask them why not! The functionality of each news program is best expalined by a local expert, I cannot possibly go into all of the details of even one news program here. Suffice to say however, if you have access to a news posting and reading program and access to the accompnaying NNTP server, you can post articles, questions and information to be read by people around the world.
Which software should I use? As I say it depends on you machine and what is available to you locally. I use a Mac and my favorite software is NewsWatcher (which is free, a definite bonus). The lastest verion of this software is available by anonymous ftp from ftp.acns.nwu.edu (the same place as Disinfectant, the antiviral utility for the Mac) in the directory /pub/newswatcher. The current version is 2.0d17. Ask around locally to see what you have available.
Why News and not just Email? The advantage of News over email mailing lists is the messages reside on a remote machine and do not clog up your mailbox. Some people have severely restricted mailbox size allocations and the output from a mailing list can swamp them and prevent them from receiving other mail. With Usenet you only download the News you want to read. You can subscribe to a small subset of the total newsgroups and selectively scan through those. You may view all of the subjects of the articles in a group before you read any of them. It is quite a flexible system.
I cant get to the Internet, what do I do? The one major complaint about Net News is that it is not accessible unless you have a connection to the Internet. The mailing lists can be accessed from Compuserve, America On-Line, GEnie, etc. An connection used to be an expensive proposition, however, these days it can be as reasonably priced as Compuserve or the other dial-up network services. For example "The World" (1-617-739-0202) charges $20 per month for 20 hours of connect time to Internet services. Another provider "The WELL" (Whole Earth 'Lectronic Link - 1-415-332-4335) charges $15 per month and $2 per hour for use. For a local provider please call the InterNIC Information Services at 1-800-444-4345.
This is just a brief outline that I have put together on the spur of the moment (as someone recently put it "its a stream of consciousness"!) Hope it helps. Drop me a line if I have made any major mistakes or left anything vital out of this message. Cheers, John Mansfield.
} 2) How it compares to laser systems? One potential problem is that to collect images using a cooled CCD camera the exposure times must be much longer than with a laser scanning confocal microscope. For instance, we did a comparison of a double labeled cultured cells scanned with a cooled CCD camera and with the BioRad MRC 600. With the CCD camera the Cy3 stain looked great (deconvolved with the quick mode of the Vaytek system). However, the FITC bleached before we could collect a few sections. On the BioRad, we were able to collect both signals simultaneously without significant bleaching of the FITC. Also, we looked at the BDS deconvolution system. We were very impressed with the deconvolved images, but here is a major difference between confocal and deconvolution: Using a confocal, investigators from all over the university can come in, scan their samples, immediately see results, get hard copy or send images to other computers instantly, and leave. If they were using deconvolution they would have to post-process every optical section before getting results. Personally, I would like to see how the BioRad confocal images would look processed with one of the deconvolution packages. -Michael Cammer
The problem "jjerome-at-isnet.is.wfu.edu (Jay Jerome)" mentioned as spectral effects when he mounted slides in glass mounts may be Newton rings. This problem may be eliminated by using anti-Newton glass in the mounts. Companies such as Wess or Gepe market this kind of glass for slide mounts. Contact your local photo shop or look in one of the photography magazines for advertisements.
} With regard to suggestion that our problem of spectral effects, we do use } Gepe anti-newton glass slides. However, the effect is reminiscent of the } effects one gets if you do not use anti-newton glass. It does not occur } however when we use thinner based film for making our slides. Only when we } use EM negative film such as 4489. Any other suggestions?
If you're not doing so already, you might try using a mask between the backing side of the film and the glass. This will eliminate the contact between the two flat surfaces that can produce Newton rings.
Way back when, we used to use commercially available masks cut from black paper. I've also used aluminum ones from EMDE Products, Inc. I don't even know whether they are still in business. The address in the literature I have from them doesn't even have a zip code. The address given is 2040 Stoner Ave., Los Angeles, CA.
John chandler-at-lamar.ColoState.EDU Fort Collins, CO
} Return-Path: {Vitor-at-dsif.fee.unicamp.br} } From: Vitor-at-dsif.fee.unicamp.br } Date: Wed, 26 Jan 94 09:26:00 EDT } To: venema-at-dutentb.et.tudelft.nl, Snitka-at-dsif.fee.unicamp.br, } tprohas-at-email.tuwien.ac.at } Subject: CALL FOR PAPERS } } } } } ----- Begin Included Message ----- } } } From PHOTOEXE%ILCTEHOL-at-VMS.HUJI.AC.IL Sun Jan 23 10:06:12 1994 } Date: Sun, 23 Jan 94 09:59 GMT } From: INTL BULLETIN ON PROCESSES AND APPLICATIONS } {PHOTOEXE%ILCTEHOL-at-VMS.HUJI.AC.IL} } Subject: CALL FOR PAPERS } To: VITOR-at-dsif.fee.unicamp.br } Content-Length: 1791 } } FRONTIERS IN SCIENCE AND TECHNOLOGY } AT NANO-MICRO SCALE } } July, 28-29 - 1994 } Guaruja - Sao Paulo State - Brazil } } Topics: } (not limited to) } } Scaling Laws - Theory } Scanning Probe Microscopies (STM, AFM,....) } Sensors } Piezoelectric Drivers } Machining (Beam, Plasma, Molecular, ...) } New Materials (Diamond-like, Porous Silicon....) } Synthesis and Structures (Biology, Microelectronics....) } Computation } ..... } } Abstract Deadline March, 20, 1994 (two printed A-4 pages) } Manuscripts at the Conference Site } } Organizing Comitttee } ==================== } } } Prof. Vitor Baranauskas - State University of Campinas - Brazil (chairperson) } Prof. Valentinas Snitka - Vibrotecnika - Lithuania } Prof. Ioshiaki Doi - State University of Campinas - Brazil } Prof. Aaron Peled - CTEH - Israel } Dr. Vladimir J. Trava-Airoldi - Instituto Nacional de Pesquisas Espaciais -Brazil } Prof. Gilberto Mattos Gualberto - State University of Campinas - Brazil } } } Organized by: } ============ } Sociedade Brasileira de Vacuo - Brazilian Vacuum Society } } This Conference will be a satellite of the Annual Brazilian Vacuum Conference, to be held in Sao Carlos City - 24-27 July, 1994 } } Guaruja is a pleasant island-city on the South Atlantic Sea. The Conference will be held in a Hotel on the beach. Proceedings will be refereed and published by an International Journal. } } For further information or abstracts submission contact : } } Prof.V.Baranauskas } Faculdade de Engenharia Eletrica } Universidade Estadual de Campinas UNICAMP } 13083-970 - Campinas - SP - Brasil } } FAX: +55-192-391395 } e.mail vitor-at-dsif.fee.unicamp.br } } ============================================================================= } ----- End Message ----- } } } } ----- End Included Message ----- } } }
Via: uk.ac.birmingham.computer-centre.ibm3090; Fri, 28 Jan 1994 11:37:44 +0000
Images from Nanoscope III can be saved as TIF files and studied on a PC using the VISILOG Software. This software is distributed (english language) by NOESIS Vision Inc 6800 Cote de Liesse Suite 200 Ville St Laurent, Quebec, Canada H4T 2A7 Tel (514) 345-1400 Fax (514) 345-1575
Fellow Microscopists: Here in the SEM lab, we scan a wide range of specimens (Insects, rodent teeth & skulls, fossil vertebrates & invertebrates) This is just to name afew. We use gold-palladium coating on these specimens. Is there anyone out there who has a sure fire method to remove coatings without damage to the specimen?
Thanks in advance, Peling Fong Melville
-------------------------------------------------------------- Peling Melville peling-at-amnh.org Interdepartmental Laboratories American Museum of Natural History
Does anyone out there have a number for Sony for color printer tech support? The loony I got in the DC area told me that I couldn't do half of the things that were printed right in the manual.
Jay: I have always made my EM slides using EM film and the only time I use glass is when I am making a composite slide. EM film mounts quite nicely in cardboard mounts, if your not doing any taping of the slides as you might do in making composite slides, why use glass? I have alwas had problems when I had to use Gepe glass. The best glass to use is by EMDE. They make an ultra thin glass (cat.# 135-NRT) slide making kit for the prevention of Newton Rings. Normal thickness glass can kill you every time with EM film. Don't know if EMDE is still in business. My box says Los Angeles 25, California. It's a few years old. You can call information in Los Angeles and see if they still have a phone # or check one of the big industrial photo supply houses or photo shop. Good Luck! Phil Rutledge prutle1-at-gl.umbc.edu
Further to Ron Anderson's suggestion: Bernie Kestel has written a paper "Non-Acid Electrolyte Thins Many Materials for TEM Without Causing Hydride Formation" (Ultramicroscopy 19 (1986) pp 205-212. I have a copy of the paper and would be pleased to FAX it to you and/or mail you a copy. You may contact me via CompuServe or as follows:
David Henriks South Bay Technology, Inc. 1120 Via Callejon San Clemente, CA 92673 USA
TEL: 714-492-2600 FAX: 714-492-1499
You may also reach Bernie Kestel at: TEL: 708-252-4945 FAX: 708-252-4798
I also have a bibliography of over 40 papers dealing primarily with TEM sample preparation. I would be pleased to send you a copy and can subsequently provide reprints at no charge.
Bernie Kestel, Argonne National Laboratory's expert at electropolishing, suggests the following for Ti alloys:
13% HCl and 87% methanol temperature = -50 C 90 Volts and 35 mA in a single jet electropolishing machine
or
60ml Percholoric Acid 590 ml Methanol 350 ml 2-butoxy ethanol (butyl cellosolve) temperature = 0 C 40-50 Volts and 50 mA in a single jet electropolishing machine
or
5.30 g LiCl 11.16 g Mg(ClO4)2 500 ml methanol 100 ml 2-butoxy ethanol (butyl cellosolve) temperature = -40 or -50 C 150 to 250 volts and 30 mA flow speed = medium single jet electropolishing machine (South Bay) The flow restriction of the Tenupol holder may require less of the 2-butoxy ethanol which increases the viscosity of the solution. Results seem less satisfactory as the temperature is raised. You may need another power supply for the higher voltage. Greater or lesser amounts may be tried of the LiCl and Mg(ClO4)2. Add the powders to the methanol with simultaneous stirring. Do not reverse the voltage polarity: this introduces hydrogen into the specimen. Keep the specimen at a positive voltage. This approach worked well on compacted Ti nanocrystalline wafers.
Message-Id: {9401282054.AA22451-at-riker.ml.wpafb.af.mil} To: WALCKSD:ml:wpafb, SCHELTFJ:ml:wpafb Subj: Electropolishing of Titanium Orig-Author: {MACLAREI-at-ibm3090.computer-centre.birmingham.ac.uk}:ddn:wpafb ----------------------------------------------------------- Dear all,
Thank you to the two of you (Rob Tayloe and Cameron Begg) who replied to my earlier message concerning the electropolishing of Titanium for TEM specimens. However I can't believe that there is no one out there that has personal experience of preparing specimens of Ti or dilute alpha-Ti alloys for the TEM. If there is anyone out there who has done some TEM on such materials I would be very grateful if you could describe the specimen preparation route that you used.
Message-Id: {9401281628.AA21291-at-riker.ml.wpafb.af.mil} To: WALCKSD:ml:wpafb, SCHELTFJ:ml:wpafb Subj: Electropolishing of Titanium Orig-Author: {MACLAREI-at-ibm3090.computer-centre.birmingham.ac.uk}:ddn:wpafb ----------------------------------------------------------- Dear all,
Thank you to the two of you (Rob Tayloe and Cameron Begg) who replied to my earlier message concerning the electropolishing of Titanium for TEM specimens. However I can't believe that there is no one out there that has personal experience of preparing specimens of Ti or dilute alpha-Ti alloys for the TEM. If there is anyone out there who has done some TEM on such materials I would be very grateful if you could describe the specimen preparation route that you used.
Message-Id: {MAILQUEUE-101.940128134052.576-at-parmly1.parmly.luc.edu} To: microscopy-at-anlemc.msd.anl.gov
} Fellow Microscopists: } Here in the SEM lab, we scan a wide range of specimens (Insects, } rodent teeth & skulls, fossil vertebrates & invertebrates) This is just to } name afew. We use gold-palladium coating on these specimens. Is there } anyone out there who has a sure fire method to remove coatings without } damage to the specimen? } } Thanks in advance, } Peling Fong Melville } } -------------------------------------------------------------- } Peling Melville peling-at-amnh.org } Interdepartmental Laboratories } American Museum of Natural History
I have never heard of such a method, and do not believe that one exists. If I'm wrong, I would be VERY interested in hearing of it. But why bother? A light coat of Au/Pd won't hurt specimens, and coated SEM specimens can be embedded for TEM or LM. If there is some compelling reason not to have a coat on your specimen, the only choice is to examine them uncoated at low kV. This is pretty simple for bones & teeth, and many inverts. It is also doable with arthropods, even allowing for their setae, but you may need to go down to 300-500 V accelerrating voltage. Phil Oshel po-at-parmly1.parmly.luc.edu
Peling: Trying to remove gold-palladium coatings can be done on specimens such as yours but it can be a pain. I have used 20-50% acetone in water and sonicated the specimens. On some specimens this has worked and on others it hasn't. I know of no sure fire method to do this. What you can use instead of gold-palladium, is silver. I have removed silver off of hard and soft tissue samples rather easily. Sometimes I need to look at cells or whole tissue by SEM and then using the same sample for TEM. I use silver most (95%) of the time. At high mags in the SEM silver has a much smaller grain size than gold or gold-palladium and I get a better signal. All it takes to remove the silver is a basic darkroom chemical called Farmers Reducing Agent. This can be bought at just about any photo shop. You just soak your sample in this solution (full strength) until the silver is removed, then do what you need to do to the sample. If I am processing for TEM, I wash the specimen 3 x 15 minutes in distilled water first, 3 x 15 minutes in buffer and start my normal processing routine for TEM starting with the dehydration cycle. If you have any questions call me or Email me. Phil Rutledge, Director Center for Electron Microscopy UMBC Bio. Dept. Phone: (410) 455-3852 FAX: (410) 455-3875 Email: prutle1-at-gl.umbc.edu
Greetings, I am looking for a postdoc to join my lab to work on the relationship between cellulose synthesis and morphogenesis. Experience with either polarized light or electron microscopy, or with spatial measurements of growth is desirable. I encourage persons of all sexes, sexual orientations, colors, nationalities, cultures, sizes, shapes and physicalities to apply. Please send me your cv, names and contact #'s (or email handles) of several references, and if available, a statement of your "world view". If you have further q's, please ask. A brief description of the project, as funded, follows.
What is the role of cellulose deposition in controlling the degree of growth anisotropy? Although it is well known that highly anisotropic growth can be rendered isotropic by the randomization of the direction of deposition of cellulose microfibrils, it is not known whether the degree of growth anisotropy is also governed by the alignment of cellulose microfibrils. The major approach taken in this project is to measure growth anisotropy at a cellular scale and then quantify the alignment of cellulose microfibrils. Cells growing at various degrees of growth anisotropy will be obtained both by taking advantage of natural variations and through use of low concentrations of microtubule inhibitors. Cellulose alignment will be quantified throughout the whole wall with polarized light microscopy and at the innermost layer in metal-carbon replicas viewed in em. Experimental material will be roots of Arabidopsis and elongating tissue culture cells. There is support to work with Prof Andrew Staehelin at Boulder to use rapid-freezing deep etch techniques to make replicas without previous fixation. The project will also focus on several root morphology mutants in arabidopsis. These appear to have tissue-specific defects in cellulose deposition, and the relation between their processing of cellulose and their aberrant morphology will be studied.
Thanks for your interest,
Tobias Baskin
******************************** *************** Tobias I. Baskin 109 Tucker Hall /~~~\ Biol. Sci's * Univ. of Missouri c|o o\ Columbia, MO 65211 USA \ = / Tel:314-882-0173 FAX 314 - 882 - 0123 """ email: baskin-at-biosci.mbp.missouri.edu
Years ago we used to study cultured hepatocytes with SEM. These cell were coated with gold/palladium. Following SEM studies we would embed these specimens in epon and section the same cells for TEM. This worked beautifully and gave the cells an interesting electron dense outline. Our SEM fix, of course, was a good glutaraldehyde/osmium fix so that the TEM fine stucture was good.
Greetings, Does anyone have any comments on the new(ish) 35 degree diamond knives that are sold by Diatome (and others?)? I am interested in comments about "ambient temperature" knives, not "cryo" knives. The Tech Staff at Diatome say that the 35 degree knives reduce compression in some samples but not all. They further said that there were NO disadvantages to the 35 degree knives. My application is with growing plant tissues, such as roots.
I am a newcomer to electron microscopy and I appreciate help from the many wise folks who are a part of this list.
Thanks in advance, Tobias Baskin
******************************** *************** Tobias I. Baskin 109 Tucker Hall /~~~\ Biol. Sci's * Univ. of Missouri c|o o\ Columbia, MO 65211 USA \ = / Tel:314-882-0173 FAX 314 - 882 - 0123 """ email: baskin-at-biosci.mbp.missouri.edu
I am investigating methods of laser illumination for light microscopy. Specifically, I would like recommendations on methods of get uniform laser illumination free of laser speckle and noise. I am aware of one reference,
Ellis, G. W., "A fiber-optic phase-randomizer for microscope illumination by laser" _J. Cell. Bio._, vol 83, p303a.
but it is rather old, 1979, and I wonder if there are better ways of doing it nowadays. It was suggested to me that a phase randomized fiber bundle might do the trick. I was wondering if anyone has tried this? Although the spatial phases of the light will then vary across the fiber bundle area, I wonder if you will still get a well defined interference pattern, which will require vibration to average away and obtain uniform illumination?
Any ideas, experiences, or references will be appreciated. And I will summarize to the list the information I receive.
Thanks, eer
Erik Reuter, e-reuter-at-uiuc.edu, 217-244-6378(W), 359-4683(H), 244-6375 fax
We are in the market to purchase some microscope objectives to operate in the near infra-red, wavelength range of 700nm to 1100nm, to be used in a lab-built microscope. We would like to achieve the highest resolution allowed by diffraction at these wavelengths (~ 1 micron?) with dry objectives.
The microscope consists of a coaxial laser illuminator, a dichroic beam splitter, an objective lens, and a eyepiece/C-mount adapter/video camera. We are examining III/V semiconductor samples.
I would like to hear your experiences and recommendations on suppliers or manufacturers of infra-red microscope objectives. Are reflecting objectives our best bet? Who makes the best reflecting objectives? In order to obtain the best performance, should we purchase the objectives together with eyepiece/C-mount adapter or camera adapters, since the objectives are probably made for a specific correction?
Thanks, eer
Erik Reuter, e-reuter-at-uiuc.edu, 217-244-6378(W), 359-4683(H), 244-6375 fax
You can get excellent illumination by using a single-mode, polarization preserving fiber coupled to an output beam expander and collimator to achieve the desired beam size for input into a CLSM for example (6 mm). BY using an adjustable collimator the gaussian beam shape is preserved at any size beam and can be adjusted to the proper back focal plane size of your objective.
All multi-mode fibers will give speckles and you have to do something to scramble this at the output (vibration, optical scramblers). See the literature on fiber optics to see why this is always going to happen to your original gaussian beam.
We have a nice fiber and input coupler from Point Source in Winchester, England FAX 44-703-602470 and are using a Spindler & Hoyer output coupler, Goettingen, FRG FAX 49-551-69-35-166.