Stain contamination and embedding in electron microscopy

1986 ◽  
Vol 44 ◽  
pp. 50-53
Author(s):  
Hilton H. Mollenhauer
Keyword(s):  
Electron Microscopy ◽  
Information Retrieval ◽  
Epoxy Resins ◽  
Image Contrast ◽  
Sample Preservation ◽  
Factors Associated ◽  
Amount Of Information ◽  
Electron Microscopical ◽  
Contrast Information

Many factors (e.g., resolution of microscope, type of tissue, and preparation of sample) affect electron microscopical images and alter the amount of information that can be retrieved from a specimen. Of interest in this report are those factors associated with the evaluation of epoxy embedded tissues. In this context, informational retrieval is dependant, in part, on the ability to “see” sample detail (e.g., contrast) and, in part, on tue quality of sample preservation. Two aspects of this problem will be discussed: 1) epoxy resins and their effect on image contrast, information retrieval, and sample preservation; and 2) the interaction between some stains commonly used for enhancing contrast and information retrieval.

Epoxy Resin Embedment

1968 ◽  
Vol 26 ◽  
pp. 100-101
Author(s):  
J. G. Adams ◽  
M. M. Campbell ◽  
H. Thomas ◽  
J. J. Ghldonl
Keyword(s):  
Electron Microscopy ◽  
Electron Beam ◽  
Epoxy Resin ◽  
Light Microscope ◽  
Epoxy Resins ◽  
Image Contrast ◽  
Tissue Preservation ◽  
Resin System ◽  
Excellent Quality

Since the introduction of epoxy resins as embedding material for electron microscopy, the list of new formulations and variations of widely accepted mixtures has grown rapidly. Described here is a resin system utilizing Maraglas 655, Dow D.E.R. 732, DDSA, and BDMA, which is a variation of the mixtures of Lockwood and Erlandson. In the development of the mixture, the Maraglas and the Dow resins were tested in 3 different volumetric proportions, 6:4, 7:3, and 8:2. Cutting qualities and characteristics of stability in the electron beam and image contrast were evaluated for these epoxy mixtures with anhydride (DDSA) to epoxy ratios of 0.4, 0.55, and 0.7. Each mixture was polymerized overnight at 60°C with 2% and 3% BDMA.Although the differences among the test resins were slight in terms of cutting ease, general tissue preservation, and stability in the beam, the 7:3 Maraglas to D.E.R. 732 ratio at an anhydride to epoxy ratio of 0.55 polymerized with 3% BDMA proved to be most consistent. The resulting plastic is relatively hard and somewhat brittle which necessitates trimming and facing the block slowly and cautiously to avoid chipping. Sections up to about 2 microns in thickness can be cut and stained with any of several light microscope stains and excellent quality light photomicrographs can be taken of such sections (Fig. 1).

Quantitative Hrem of Electron-Irradiated Quartz

1994 ◽  
Vol 373 ◽  
Author(s):  
L.C. Qin ◽  
L.W. Hobbs
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Morphological Evolution ◽  
High Resolution Electron Microscopy ◽  
Image Contrast ◽  
Total Thickness ◽  
Resolution Electron ◽  
Electron Microscopical ◽  
Transformation Front

AbstractThe progress of heterogeneous amorphization of electron-irradiated quartz was studied in situ by high-resolution electron microscopy (HREM). The transformation front appears surprisingly sharp when viewed close to edge-on, but the apparent morphological evolution when aperiodic and periodic material overlap in projection is one in which gradual changes in image regularity are observed. Electron-microscopical contrast obtained in HREM irradiations was interpreted quantitatively, through the use of image contrast simulations. When the amorphized portion occupies less than about two-thirds of the total thickness, the HREM images continue to appear crystalline. Only when the amorphized thickness fraction exceeds about two-thirds does the resultant image contrast show mainly non-crystalline features.

Observability of Very Thick Specimen by Using Transmission Scanning Electron Microscope

1971 ◽  
Vol 29 ◽  
pp. 26-27
Author(s):  
K. Shibatomi ◽  
T. Yamanoto ◽  
H. Koike
Keyword(s):  
Electron Microscope ◽  
Image Quality ◽  
Scanning Electron Microscope ◽  
Chromatic Aberration ◽  
Image Contrast ◽  
Objective Lens ◽  
Thick Specimen ◽  
Transmission Electron ◽  
Scanning Electron

In the observation of a thick specimen by means of a transmission electron microscope, the intensity of electrons passing through the objective lens aperture is greatly reduced. So that the image is almost invisible. In addition to this fact, it have been reported that a chromatic aberration causes the deterioration of the image contrast rather than that of the resolution. The scanning electron microscope is, however, capable of electrically amplifying the signal of the decreasing intensity, and also free from a chromatic aberration so that the deterioration of the image contrast due to the aberration can be prevented. The electrical improvement of the image quality can be carried out by using the fascionating features of the SEM, that is, the amplification of a weak in-put signal forming the image and the descriminating action of the heigh level signal of the background. This paper reports some of the experimental results about the thickness dependence of the observability and quality of the image in the case of the transmission SEM.

The Current Situation in Chemical Stabilization of Biological Materials

1972 ◽  
Vol 30 ◽  
pp. 132-133
Author(s):  
John H. Luft
Keyword(s):  
Electron Microscopy ◽  
Electron Microscope ◽  
Osmium Tetroxide ◽  
Molecular Level ◽  
Cross Linking ◽  
Chemical Stabilization ◽  
Specimen Preparation ◽  
Sufficient Condition ◽  
Radio Telescopes

With information processing devices such as radio telescopes, microscopes or hi-fi systems, the quality of the output often is limited by distortion or noise introduced at the input stage of the device. This analogy can be extended usefully to specimen preparation for the electron microscope; fixation, which initiates the processing sequence, is the single most important step and, unfortunately, is the least well understood. Although there is an abundance of fixation mixtures recommended in the light microscopy literature, osmium tetroxide and glutaraldehyde are favored for electron microscopy. These fixatives react vigorously with proteins at the molecular level. There is clear evidence for the cross-linking of proteins both by osmium tetroxide and glutaraldehyde and cross-linking may be a necessary if not sufficient condition to define fixatives as a class.

A light optical diffractometer for electron microscopical images operating in line

1978 ◽  
Vol 36 (1) ◽  
pp. 86-87
Author(s):  
P. Bonhomme ◽  
A. Beorchia
Keyword(s):  
Electron Microscopy ◽  
Electron Transfer ◽  
Electron Microscope ◽  
Image Converter ◽  
Coherent Light ◽  
Spatial Filters ◽  
Electron Image ◽  
Electron Microscopical ◽  
Working Parameters ◽  
Amorphous Part

We have already described (1.2.3) a device using a pockel's effect light valve as a microscopical electron image converter. This converter can be read out with incoherent or coherent light. In the last case we can set in line with the converter an optical diffractometer. Now, electron microscopy developments have pointed out different advantages of diffractometry. Indeed diffractogram of an image of a thin amorphous part of a specimen gives information about electron transfer function and a single look at a diffractogram informs on focus, drift, residual astigmatism, and after standardizing, on periods resolved (4.5.6). These informations are obvious from diffractogram but are usualy obtained from a micrograph, so that a correction of electron microscope parameters cannot be realized before recording the micrograph. Diffractometer allows also processing of images by setting spatial filters in diffractogram plane (7) or by reconstruction of Fraunhofer image (8). Using Electrotitus read out with coherent light and fitted to a diffractometer; all these possibilities may be realized in pseudoreal time, so that working parameters may be optimally adjusted before recording a micrograph or before processing an image.

Ultrastructure of the soil fungus, Geomyces pannorus

1984 ◽  
Vol 42 ◽  
pp. 738-739
Author(s):  
J. A. Traquair ◽  
E. G. Kokko
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Fine Structure ◽  
Low Temperatures ◽  
Soil Fungus ◽  
Organic Soils ◽  
Anatomical Studies ◽  
Transmission Electron ◽  
Electron Microscopical ◽  
Scanning Electron

With the advent of improved dehydration techniques, scanning electron microscopy has become routine in anatomical studies of fungi. Fine structure of hyphae and spore surfaces has been illustrated for many hyphomycetes, and yet, the ultrastructure of the ubiquitous soil fungus, Geomyces pannorus (Link) Sigler & Carmichael has been neglected. This presentation shows that scanning and transmission electron microscopical data must be correlated in resolving septal structure and conidial release in G. pannorus.Although it is reported to be cellulolytic but not keratinolytic, G. pannorus is found on human skin, animals, birds, mushrooms, dung, roots, and frozen meat in addition to various organic soils. In fact, it readily adapts to growth at low temperatures.

Characterization of homoepitaxial diamond films by Electron microscopy

1991 ◽  
Vol 49 ◽  
pp. 880-881
Author(s):  
D.P. Malta ◽  
S.A. Willard ◽  
R.A. Rudder ◽  
G.C. Hudson ◽  
J.B. Posthill ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Surface Defects ◽  
Substrate Surface ◽  
Diamond Films ◽  
Chemical Vapor ◽  
Polycrystalline Diamond ◽  
Large Degree ◽  
Rf Plasma ◽  
Semiconducting Diamond

Semiconducting diamond films have the potential for use as a material in which to build active electronic devices capable of operating at high temperatures or in high radiation environments. A major goal of current device-related diamond research is to achieve a high quality epitaxial film on an inexpensive, readily available, non-native substrate. One step in the process of achieving this goal is understanding the nucleation and growth processes of diamond films on diamond substrates. Electron microscopy has already proven invaluable for assessing polycrystalline diamond films grown on nonnative surfaces.The quality of the grown diamond film depends on several factors, one of which is the quality of the diamond substrate. Substrates commercially available today have often been found to have scratched surfaces resulting from the polishing process (Fig. 1a). Electron beam-induced current (EBIC) imaging shows that electrically active sub-surface defects can be present to a large degree (Fig. 1c). Growth of homoepitaxial diamond films by rf plasma-enhanced chemical vapor deposition (PECVD) has been found to planarize the scratched substrate surface (Fig. 1b).

The annealed (0001) α-alumina surface and its influence on thin-film growth

1995 ◽  
Vol 53 ◽  
pp. 336-337
Author(s):  
Michael W. Bench ◽  
Paul G. Kotula ◽  
C. Barry Carter
Keyword(s):  
Electron Microscopy ◽  
Surface Energy ◽  
Film Growth ◽  
Thin Film Growth ◽  
Energy Calculations ◽  
Transmission Electron ◽  
Reflection Electron Microscopy ◽  
Low Energy Electron ◽  
Surface Nature

The growth of semiconductors, superconductors, metals, and other insulators has been investigated using alumina substrates in a variety of orientations. The surface state of the alumina (for example surface reconstruction and step nature) can be expected to affect the growth nature and quality of the epilayers. As such, the surface nature has been studied using a number of techniques including low energy electron diffraction (LEED), reflection electron microscopy (REM), transmission electron microscopy (TEM), molecular dynamics computer simulations, and also by theoretical surface energy calculations. In the (0001) orientation, the bulk alumina lattice can be thought of as a layered structure with A1-A1-O stacking. This gives three possible terminations of the bulk alumina lattice, with theoretical surface energy calculations suggesting that termination should occur between the Al layers. Thus, the lattice often has been described as being made up of layers of (Al-O-Al) unit stacking sequences. There is a 180° rotation in the surface symmetry of successive layers and a total of six layers are required to form the alumina unit cell.

Technical notes on routine TEM procedures

1987 ◽  
Vol 45 ◽  
pp. 630-631
Author(s):  
K. Chien ◽  
R.L. Van de Velde ◽  
R.C. Heusser
Keyword(s):  
Epoxy Resins ◽  
Pathology Laboratory ◽  
Silicone Fluid ◽  
Image Position

Sectioning quality of epoxy resins can be improved by the addition of a 1% silicone 200 fluid (Dow Corning), however this produces a softer block. To compensate, a harder plastic has been used for embedding various tissues encountered in our pathology laboratory. Exact amounts of the plastic mixture can be directly made up for embedding as shown: The chart reveals a Poly/Bed 812 (WPE 145) to anhydride ratio of 1:0.7 and a NMA to DDSA ratio of 7:3. 1% silicone fluid is added to above mixtures.Due to impurities within the DDSA and NMA, the polymerized epoxy blocks vary in darkness and appear to affect sectioning quality. After discussing this problem with Polysciences Inc., they have agreed to purify their anhydrides in an effort to standardize the consistency of the plastic.

Methods to Monitor and Improve the Performance of Specimen Holders for Transmission Electron Cryomicroscopy

1996 ◽  
Vol 54 ◽  
pp. 454-455
Author(s):  
B. L. Armbruster ◽  
B. Kraus ◽  
M. Pan
Keyword(s):  
Electron Microscopy ◽  
Electron Beam ◽  
Electron Beam Irradiation ◽  
Beam Irradiation ◽  
Quality Of Data ◽  
Electron Cryomicroscopy ◽  
Thermal Equilibration ◽  
Transmission Electron ◽  
Electron Microscopes

One goal in electron microscopy of biological specimens is to improve the quality of data to equal the resolution capabilities of modem transmission electron microscopes. Radiation damage and beam- induced movement caused by charging of the sample, low image contrast at high resolution, and sensitivity to external vibration and drift in side entry specimen holders limit the effective resolution one can achieve. Several methods have been developed to address these limitations: cryomethods are widely employed to preserve and stabilize specimens against some of the adverse effects of the vacuum and electron beam irradiation, spot-scan imaging reduces charging and associated beam-induced movement, and energy-filtered imaging removes the “fog” caused by inelastic scattering of electrons which is particularly pronounced in thick specimens.Although most cryoholders can easily achieve a 3.4Å resolution specification, information perpendicular to the goniometer axis may be degraded due to vibration. Absolute drift after mechanical and thermal equilibration as well as drift after movement of a holder may cause loss of resolution in any direction.

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