Preparatory Procedures for Electron Microscopic Analysis at the Molecular Level of Resolution

1978 ◽  
Vol 36 (3) ◽  
pp. 516-520
Author(s):  
F.J. Sjostrand
Keyword(s):  
Electron Microscope ◽  
Molecular Level ◽  
Microscopic Analysis ◽  
Resolving Power ◽  
Cellular Structures ◽  
Electron Microscopic ◽  
Systematic Analysis ◽  
Technical Developments ◽  
Thin Sectioning ◽  
Obvious Reason

In the 1940's and 1950's electron microscopy conferences were attended with everybody interested in learning about the latest technical developments for one very obvious reason. There was the electron microscope with its outstanding performance but nobody could make very much use of it because we were lacking proper techniques to prepare biological specimens. The development of the thin sectioning technique with its perfectioning in 1952 changed the situation and systematic analysis of the structure of cells could now be pursued. Since then electron microscopists have in general become satisfied with the level of resolution at which cellular structures can be analyzed when applying this technique. There has been little interest in trying to push the limit of resolution closer to that determined by the resolving power of the electron microscope.

Bridging the Resolution Gap: Correlated 3D Light and Electron Microscopic Analysis of Large Biological Structures

1999 ◽  
Vol 5 (S2) ◽  
pp. 526-527
Author(s):  
Maryann E. Martone
Keyword(s):  
Electron Microscopy ◽  
Electron Microscope ◽  
Surface Area ◽  
Light Microscope ◽  
Microscopic Analysis ◽  
Electron Microscopic Analysis ◽  
Electron Microscopic ◽  
Transmission Electron ◽  
Biological Structures ◽  
Ganglion Neurons

One class of biological structures that has always presented special difficulties to scientists interested in quantitative analysis is comprised of extended structures that possess fine structural features. Examples of these structures include neuronal spiny dendrites and organelles such as the Golgi apparatus and endoplasmic reticulum. Such structures may extend 10's or even 100's of microns, a size range best visualized with the light microscope, yet possess fine structural detail on the order of nanometers that require the electron microscope to resolve. Quantitative information, such as surface area, volume and the micro-distribution of cellular constituents, is often required for the development of accurate structural models of cells and organelle systems and for assessing and characterizing changes due to experimental manipulation. Performing estimates of such quantities from light microscopic data can result in gross inaccuracies because the contribution to total morphometries of delicate features such as membrane undulations and excrescences can be quite significant. For example, in a recent study by Shoop et al, electron microscopic analysis of cultured chick ciliary ganglion neurons showed that spiny projections from the plasmalemma that were not well resolved in the light microscope effectively doubled the surface area of these neurons.While the resolution provided by the electron microscope has yet to be matched or replaced by light microscopic methods, one drawback of electron microscopic analysis has always been the relatively small sample size and limited 3D information that can be obtained from samples prepared for conventional transmission electron microscopy. Reconstruction from serial electron micrographs has provided one way to circumvent this latter problem, but remains one of the most technically demanding skills in electron microscopy. Another approach to 3D electron microscopic imaging is high voltage electron microscopy (HVEM). The greater accelerating voltages of HVEM's allows for the use of much thicker specimens than conventional transmission electron microscopes.

Electron microscope studies of the isolated dna fragments containing biotin genes of escherichia coli k12

1978 ◽  
Vol 36 (2) ◽  
pp. 198-199
Author(s):  
C.K. DasGupta ◽  
H. Samanta ◽  
A. Guha
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Electron Microscope ◽  
Linkage Map ◽  
Molecular Level ◽  
Present Report ◽  
Biosynthetic Enzymes ◽  
Electron Microscopic ◽  
Phage Dna

The genes coding for the biotin biosynthetic enzymes, bioABFCD are located at 17 min of the linkage map of EL coli K12 (Bachman, et al. Bact. Rev. 40: 116, 1976). It was shown that the bioA gene is oriented anticlockwise and the bioBFCD genes are oriented clockwise in the EL coli map, with the bio regulator region lying in between them (Guha et al. J. Mol. Biol. 56: 53, 1971). The study of the control elements of this divergently oriented operon at the molecular level is important for understanding the various intriguing mechanisms of gene expression in bacteria. In the present work, the electron microscopic characterization of the entire biotin genes and its regulator segment isolated after cleaving a biotin transducing phage DNA by restriction endonucleases, is described. The isolated biotin DNA fratments were subsequently recombined with the colEI DNA to isolate a colEl-biotin DNA hybrid. The present report also includes a portion of the study of this cloned plasmid chimera in a biotin auxotroph.

Thrombin and Reptilase Induced Fibrin Structures in the Presence of Dextran: A Quantitative Electron Microscope Study

1975 ◽  
Author(s):  
W. H. Krause ◽  
P. Zimmermann
Keyword(s):  
Electron Microscope ◽  
Microscope Study ◽  
Electron Microscope Study ◽  
Microscopic Analysis ◽  
Electron Microscopic Analysis ◽  
Segment Length ◽  
Electron Microscopic ◽  
Fibrin Network ◽  
Significant Difference ◽  
Microscopic Studies

The present study describes an electron microscopic analysis, using a quantitative morphometric method, of fibrin network obtained by thrombin and reptilase in the presence of dextran. The thickness of the thrombin induced fibrin meshwork show a significant difference compared to fibrin formed by reptilase. The segment length of reptilase fibrin are reduced to 25% and the thickness of the fibers are reduced to 75% in comparison with thrombin fibrin.The thickness of the fibrin meshwork obtained by thrombin and/or reptilase in the presence of dextran ( w 40,000) is significantly diminished compared to saline controls (p < 0.001). Segment length and thickness of fibers in the clots with dextran showed different results when thrombin or reptilase were used as enzyme.The investigation indicates that the fibrin meshwork formed by thrombin and/or reptilase in the presence or absence of dextran result in a significantly different fibrin morphylogy. The results are different to the so far descriptive electron microscopic studies.

Somitogenesis in the amphibian Xenopus laevis: scanning electron microscopic analysis of intrasomitic cellular arrangements during somite rotation

Development ◽  
1981 ◽  
Vol 64 (1) ◽  
pp. 23-43
Author(s):  
B. Woo Youn ◽  
George M. Malacinski
Keyword(s):  
Electron Microscope ◽  
Scanning Electron Microscope ◽  
Xenopus Laevis ◽  
Microscopic Analysis ◽  
Block Rotation ◽  
Electron Microscopic ◽  
Cellular Changes ◽  
Scanning Electron Microscopic ◽  
Scanning Electron Microscopic Analysis ◽  
Scanning Electron

The intrasomitic changes in cell arrangement which accompany somite rotation during somitogenesis in Xenopus laevis were analysed with the scanning electron microscope (SEM). Longitudinal, horizontal fractures of whole embryos were examined at various dorsoventral levels of stage-22 to -24 embryos. Observations of the gross morphological features of somitogenesis, and the cellular changes which accompany somite segmentation and somite rotation were made. Several of these observations lead to modifications of previous models for the cellular basis of somitogenesis in Xenopus. Individual cellular rearrangements, rather than simultaneous block rotation of a whole somite, appear to be responsible for the 90° rotation of myotomal cells within a single somite. Cellular arrangments in fused somites were also examined. Some ultraviolet-irradiated embryos displayed a complete lack of a notochord. The somites in those embryos were fused across the midline beneath the neural tube. The dorsal and ventral arms of the somites are not fused. Normal rotation occurs only in the dorsal and ventral arms while, in the majority of cases, cells in the fused region fail to rotate normally. In some cases, individual cells in the fused region undergo partial rearrangement. Those observations support the notion that individual cellular rearrangements account for the rotation of the whole somite.

scholarly journals Viroplasm matrix protein Pns9 from rice gall dwarf virus forms an octameric cylindrical structure

2011 ◽  
Vol 92 (9) ◽  
pp. 2214-2221 ◽  
Author(s):  
Fusamichi Akita ◽  
Naoyuki Miyazaki ◽  
Hiroyuki Hibino ◽  
Takumi Shimizu ◽  
Akifumi Higashiura ◽  
...  
Keyword(s):  
Electron Microscope ◽  
Matrix Protein ◽  
Three Dimensional ◽  
Microscopic Analysis ◽  
Host Cells ◽  
Size Exclusion ◽  
Dwarf Virus ◽  
Electron Microscopic ◽  
The Family ◽  
Rice Gall Dwarf Virus

The non-structural Pns9 protein of rice gall dwarf virus (RGDV) accumulates in viroplasm inclusions, which are structures that appear to play an important role in viral morphogenesis and are commonly found in host cells infected by viruses in the family Reoviridae. Immunofluorescence and immunoelectron microscopy of RGDV-infected vector cells in monolayers, using antibodies against Pns9 of RGDV and expression of Pns9 in Spodoptera frugiperda cells, demonstrated that Pns9 is the minimal viral factor necessary for formation of viroplasm inclusion during infection by RGDV. When Pns9 in solution was observed under a conventional electron microscope, it appeared as ring-like aggregates of approximately 100 Å in diameter. Cryo-electron microscopic analysis of these aggregates revealed cylinders of octameric Pns9, whose dimensions were similar to those observed under the conventional electron microscope. Octamerization of Pns9 in solution was confirmed by the results of size-exclusion chromatography. Among proteins of viruses that belong to the family Reoviridae whose three-dimensional structures are available, a matrix protein of the viroplasm of rotavirus, NSP2, forms similar octamers, an observation that suggests similar roles for Pns9 and NSP2 in morphogenesis in animal-infecting and in plant-infecting reoviruses.

scholarly journals An Electron Microscope Study of Sectioned Cells of Peripheral Blood and Bone Marrow

Blood ◽  
1954 ◽  
Vol 9 (1) ◽  
pp. 24-38 ◽  
Author(s):  
JEAN KAUTZ ◽  
Q. B. DEMARSH
Keyword(s):  
Bone Marrow ◽  
Fine Structure ◽  
Electron Microscope ◽  
Peripheral Blood ◽  
Blood Cells ◽  
Electron Microscope Study ◽  
White Blood Cells ◽  
Cell Types ◽  
Resolving Power ◽  
Thin Sectioning

Abstract White blood cells from peripheral blood and bone marrow have been sectioned for study in the electron microscope. Methods of fixation and handling are described. Most of the usual blood cell types have been tentatively identified, and their fine structure is described. The high resolving power of the electron microscope promises to reveal details previously unsuspected, as well as to extend and clarify existing knowledge concerning the cytology of blood cells, both normal and pathologic. Ultra-thin sectioning, while still a very difficult art, appears to be the best method currently available for visualizing the fine structure of white blood cells, which would otherwise be too thick for penetration by the electron beam. Conditions of satisfactory fixation and dehydration are extremely critical, and care must be exercised in the interpretation of all results in order to separate gross fixation artifacts from the finer precipitation of protoplasmic material which may approximate a true picture of the living cell.

A Reproducible Selective Stain for Cardiac Sarcoplasmic Reticulum

1974 ◽  
Vol 32 ◽  
pp. 214-215
Author(s):  
R. A. Waugh ◽  
J. R. Sommer
Keyword(s):  
Complex System ◽  
Electron Microscope ◽  
Sarcoplasmic Reticulum ◽  
Transmission Electron Microscope ◽  
Electron Microscopic ◽  
Cardiac Sarcoplasmic Reticulum ◽  
Transmission Electron

Cardiac sarcoplasmic reticulum (SR) is a complex system of intracellular tubules that, due to their small size and juxtaposition to such electron-dense structures as mitochondria and myofibrils, are often inconspicuous in conventionally prepared electron microscopic material. This study reports a method with which the SR is selectively “stained” which facilitates visualizationwith the transmission electron microscope.

Observation of biological macromolecules using various electron microscopic methods

1978 ◽  
Vol 36 (2) ◽  
pp. 170-171
Author(s):  
Mitsuo Ohtsuki ◽  
Michael Sogard
Keyword(s):  
Electron Microscope ◽  
Phase Contrast ◽  
Image Interpretation ◽  
Density Lipoprotein ◽  
Biological Macromolecules ◽  
Contrast Effects ◽  
Biological Structure ◽  
Electron Microscopic ◽  
Structural Investigations ◽  
Staining Techniques

Structural investigations of biological macromolecules commonly employ CTEM with negative staining techniques. Difficulties in valid image interpretation arise, however, due to problems such as variability in thickness and degree of penetration of the staining agent, noise from the supporting film, and artifacts from defocus phase contrast effects. In order to determine the effects of these variables on biological structure, as seen by the electron microscope, negative stained macromolecules of high density lipoprotein-3 (HDL3) from human serum were analyzed with both CTEM and STEM, and results were then compared with CTEM micrographs of freeze-etched HDL3. In addition, we altered the structure of this molecule by digesting away its phospholipid component with phospholipase A2 and look for consistent changes in structure.

The High Resolution Scanning Transmission Electron Microscope

1974 ◽  
Vol 32 ◽  
pp. 316-317
Author(s):  
A. V. Crewe
Keyword(s):  
Electron Microscope ◽  
High Resolution ◽  
High Vacuum ◽  
Scanning Transmission Electron Microscope ◽  
Ultra High Vacuum ◽  
Resolving Power ◽  
Imaging Characteristics ◽  
Transmission Electron ◽  
Scanning Transmission ◽  
The Moment

The high resolution STEM is now a fact of life. I think that we have, in the last few years, demonstrated that this instrument is capable of the same resolving power as a CEM but is sufficiently different in its imaging characteristics to offer some real advantages.It seems possible to prove in a quite general way that only a field emission source can give adequate intensity for the highest resolution^ and at the moment this means operating at ultra high vacuum levels. Our experience, however, is that neither the source nor the vacuum are difficult to manage and indeed are simpler than many other systems and substantially trouble-free.

Transmission Electron Microscopy of Protein Macromolecules

1971 ◽  
Vol 29 ◽  
pp. 424-425
Author(s):  
Henry S. Slayter
Keyword(s):  
Biological Systems ◽  
Metal Vapor ◽  
Resolving Power ◽  
Electron Microscopic ◽  
Chemical Methods ◽  
Molecular Weights ◽  
The Past ◽  
Physical Chemical ◽  
Transmission Electron

Electron microscopic methods have been applied increasingly during the past fifteen years, to problems in structural molecular biology. Used in conjunction with physical chemical methods and/or Fourier methods of analysis, they constitute powerful tools for determining sizes, shapes and modes of aggregation of biopolymers with molecular weights greater than 50, 000. However, the application of the e.m. to the determination of very fine structure approaching the limit of instrumental resolving power in biological systems has not been productive, due to various difficulties such as the destructive effects of dehydration, damage to the specimen by the electron beam, and lack of adequate and specific contrast. One of the most satisfactory methods for contrasting individual macromolecules involves the deposition of heavy metal vapor upon the specimen. We have investigated this process, and present here what we believe to be the more important considerations for optimizing it. Results of the application of these methods to several biological systems including muscle proteins, fibrinogen, ribosomes and chromatin will be discussed.

Sign in / Sign up

Export Citation Format

Share Document