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PubMed1975

[Scanning transmission electron microscopy of dendritic spines stained by the Golgi method].

Scanning transmission electron microscopy of the dendritic spines of multipolar neurons in the cat inferior Colliculus was achieved on Golgi semi-thin sections. The three basic types of dendritic spines (ST, MS, TH) were identified. Scanning transmission electron microscopy provides a reliable method for a three dimensional view of these structures at high resolution and consequently a more accurate appreciation of their size. In addition, it could prove very useful in the quantitative analysis of the dendritic spines.

Animals↗
PubMed1997

Phosphorylation and subunit organization of axonal neurofilaments determined by scanning transmission electron microscopy.

Phosphorylation plays a critical role in controlling the function of cytoskeletal assemblies but no direct method yet exists to measure the phosphorylation state of proteins at the level of individual molecules and assemblies. Herein, we apply scanning transmission electron microscopy in combination with electron energy loss spectroscopy to measure the distributions of mass and phosphorus in neurofilaments (NFs) isolated from the squid giant axon. We find that native squid NFs, in contrast to typical reconstituted intermediate filaments, are a relatively homogeneous population containing only eight coiled-coil dimers per cross section. The measured stoichiometry of approximately 1:1 for light/heavy peptides strongly suggests that squid NFs are composed of heterodimers. Furthermore, each heavy chain of the dimers carries at least 100 phosphate groups and is, therefore, near-maximally phosphorylated. These results also demonstrate that scanning transmission electron microscopy combined with electron energy loss spectroscopy at the nanometer scale is capable of characterizing the level and distribution of phosphorylation in individual mass-mapped protein assemblies.

Animals↗
PubMed1983

Visualization of anti-histone antibodies on SV40 minichromosomes by scanning transmission electron microscopy (STEM).

The unique capabilities of the scanning transmission electron microscope (STEM) have been used for a high resolution study of antibody binding to individual SV40 minichromosomes. A method of sample preparation has been developed which allows direct visualization of the antibody molecules in a clearly recognizable form. Using this technique, we have studied the binding of anti-H2B and anti-H3 immunoglobulins to SV40 minichromosomes. The results indicate that histones H2B and H3 are located only in the nucleosomes and are absent in the linker regions.

Animals↗
PubMed1988

Titin: quantitative mass measurements by scanning transmission electron microscopy and structural implications for the sarcomere matrix of skeletal muscle.

Scanning transmission electron microscopy has been used to investigate mass and linear mass density of native titin-2, a large soluble fragment of intact titin, from rabbit skeletal muscle. Dark field images of unstained, freeze-dried titin-2 appeared as either compact globules or looser and larger balls of string. Direct mass measurements indicated that the compact forms have an average mass of 2.40 +/- 0.50 x 10(6) Da. The mass to length ratio, determined from well-spread portions of titin strands (3-5 nm wide) from the ball of string forms, averaged 2.7 +/- 0.9 kDa/nm. Thus a single native intact titin molecule has a calculated contour length of well above approximately 1 micron, sufficient to span unidirectionally between the Z line and M line region in a resting-length sarcomere.

Animals↗
PubMedDate not supplied

Heterogeneity of Escherichia coli ribosomes established by scanning transmission electron microscopy.

Quantitative mass image analysis of Escherichia coli ribosomal particles by scanning transmission electron microscopy (STEM) provided direct evidence that presumably homogeneous preparations of ribosomes are, in reality, populations of heterogeneous particles. Variations in composition, relative molecular mass (Mr) and shape were observed both in the monosomes and in the ribosomal subunits. None of these changes can be resolved visually; they can be evaluated only by computer processing. The variations in relative mass and shape monitored by values of radius of gyration (RG) were attributed to the loss of ribosomal proteins and/or factors and correlated with the changes in ribosome composition and biological activity. The highest activity was found in monosomes prepared from the standard 0.5 M NH4Cl wash. With increasing concentrations (up to 1.5 M) of NH4Cl in the wash buffer the activity decreased slowly, then dropped rapidly to about half in 2 M NH4Cl. The most striking effects were observed in ribosomal particles washed with 0.1 M NH4Cl. The 70S monosomes and the 30S subunits attained maximum Mr and RG values (2660 kDa and 76 A, and 990 kDa and 75 A, respectively), which were greater than the theoretical values, while the activity was minimal (approximately 12%). The Mr and RG parameters of the 50S subunits remained uneffected by the NH4Cl washes (approximately 1600 kDa and 68 A).

Escherichia coli↗
PubMed2005

Valence electron energy-loss spectroscopy in monochromated scanning transmission electron microscopy.

With the development of monochromators for (scanning) transmission electron microscopes, valence electron energy-loss spectroscopy (VEELS) is developing into a unique technique to study the band structure and optical properties of nanoscale materials. This article discusses practical aspects of spatially resolved VEELS performed in scanning transmission mode and the alignments necessary to achieve the current optimum performance of approximately 0.15 eV energy resolution with an electron probe size of approximately 1 nm. In particular, a collection of basic concepts concerning the acquisition process, the optimization of the energy resolution, the spatial resolution and the data processing are provided. A brief study of planar defects in a Y(1)Ba(2)Cu(3)O(7-)(delta) high-temperature superconductor illustrates these concepts and shows what kind of information can be accessed by VEELS.

Journal Article↗
PubMed1991

Mass analysis of bacteriophage T4 proheads and mature heads by scanning transmission electron microscopy and hydrodynamic measurements.

Quantitative mass analysis of bacteriophage T4 proheads by scanning transmission electron microscopy (STEM) revealed a mass of 79.5 +/- 0.6 MDa, while hydrodynamic measurements yielded a prohead mass of about 80 MDa. This is 25% less than the prohead mass deduced from its polypeptide composition, and this finding implies that the bacteriophage T4 prohead is built of fewer polypeptide copies than previously reported. In contrast, the mass of mature heads measured by STEM, 194 +/- 2 MDa, is in agreement with previous mass measurements of DNA and protein content, and it is consistent with the previously determined stoichiometry. This good agreement of average STEM values for proheads and mature heads with corresponding hydrodynamic measurements suggests that STEM allows faithful evaluation of the masses of large supramolecular assemblies (i.e., greater than or equal to 200 MDa) such as whole viruses or cellular organelles.

Formaldehyde↗
PubMed1985

Studies on the structure of bovine factor V by scanning transmission electron microscopy.

We studied purified bovine factor V (mol wt 330,000) by scanning transmission electron microscopy (STEM) of freeze-dried unstained or negatively contrasted preparations. Freeze-dried molecules revealed discrete shapes ranging from roughly spheroidal (100 to 120 nm) to oblong (140 to 200 nm in length X 50 to 100 nm in width). Oblong shapes could often be resolved into two or three distinct domains, ranging from 60 to 100 nm in diameter. A "satellite" nodular structure (30 to 50 nm in diameter) connected to the main molecule by a thin stalk (approximately 10 nm wide) up to 80 nm in length was occasionally seen. Glutaraldehyde-treated preparations yielded the same shapes as were seen in unfixed preparations but revealed better definition of submolecular features and "satellite" nodules. STEM mass analysis confirmed that each of the different shapes represented a monomolecular form of factor V. Negatively stained images revealed objects having the same general shapes as freeze-dried molecules, although greater detail was evident. Some images suggested that molecules consist of five or more discrete parts. Taken together, these observations indicate that factor V molecules are multidomainal, flexible structures that tend to have an irregular oblong shape with an axial ratio between 3:2 and 2:1.

Animals↗
PubMed2005

Scanning transmission electron microscopy and its application to the study of nanoparticles and nanoparticle systems.

Scanning transmission electron microscopy (STEM) techniques can provide imaging, diffraction and spectroscopic information, either simultaneously or in a serial manner, of the specimen with an atomic or a sub-nanometer spatial resolution. High-resolution STEM imaging, when combined with nanodiffraction, atomic resolution electron energy-loss spectroscopy and nanometer resolution X-ray energy dispersive spectroscopy techniques, is critical to the fundamental studies of importance to nanoscience and nanotechnology. The availability of sub-nanometer or sub-angstrom electron probes in a STEM instrument, due to the use of a field emission gun and aberration correctors, ensures the greatest capabilities for studies of sizes, shapes, defects, crystal and surface structures, and compositions and electronic states of nanometer-size regions of thin films, nanoparticles and nanoparticle systems. The various imaging, diffraction and spectroscopy modes available in a dedicated STEM or a field emission TEM/STEM instrument are reviewed and the application of these techniques to the study of nanoparticles and nanostructured catalysts is used as an example to illustrate the critical role of the various STEM techniques in nanotechnology and nanoscience research.

Journal Article↗
PubMed2000

Simulation of atomic-scale high-angle annular dark field scanning transmission electron microscopy images.

Image simulations for high-angle annular dark field (HAADF) scanning transmission electron microscopy (STEM) based on the Bethe's eigen-value method are presented. The effects of aperture size and defocus of a probe-forming lens, both of which determine the shape of the probe, and the effect of the distortion, influencing the Bloch wave field on atomic columns channelled by electrons, on the HAADF-image intensity are discussed in terms of dynamical effect. These effects are illustrated by our experimental atomic-scale HAADF-STEM images, detected in a detector range of 50-110 mrad. It is emphasized that the image simulations are indispensable for quantification of experimental HAADF-STEM images and as such provide a valuable compositional analysis for every atomic column along the incident beam.

Journal Article↗
PubMed1991

Morphologic changes in the human amnion epithelium that accompany labor as seen with scanning and transmission electron microscopy.

Scanning and transmission electron microscopy were used to assess the influence of normal, active labor on the ultrastructure of the human amnion epithelial membrane. Amnion membranes (reflected and placental portions) were obtained from patients either in active labor who were delivered vaginally or by cesarean section after 6 to 12 hours of labor or from patients who underwent elective cesarean section before clinical signs of overt labor. Scanning electron microscopy revealed that reflected amnion membranes that were obtained from patients who were not in labor consisted of a uniform single layer of epithelial cells with numerous microvilli on the apical surface and closely associated cellular borders. In contrast, amnion membranes that were obtained from patients who were in labor consisted of a single layer of epithelial cells, which was interrupted by wide intercellular gaps and extracellular extrusions. Transmission electron microscopy showed that intercellular junctions tended to be less complex in patients who were in labor versus patients who were not in labor. Although lipid droplets were prevalent in both patient groups, specimens that were obtained from patients who were in labor had more lipid droplets per cell than specimens from patients who were not in labor. These results support the theory that the complex biochemical events that culminate in parturition are accompanied and/or preceded by demonstrable morphologic changes in the amnion membrane.

Amnion↗
PubMed1975

Electron microscopic studies of the lung of the frog. II. Topography of the inner surface by scanning and transmission electron microscopy.

Scanning and transmission electron microscopy were used to study the inner architecture of the frog lung. In some specimens the alveolar surface mucus layer was removed to permit the examination of underlying features. The inner surface of the frog's lung is covered by a layer of microvilli belonging to only one type of epithelial cells. The boundaries of these epithelial cells are demarcated by small ridges. Different degrees of lung expansion cause variations of the surface topography. The morphology of certain surface features is examined in detail. Several methods of drying the specimens are compared.

Animals↗
PubMed2004

Low-voltage electron microscopy of polymer and organic molecular thin films.

We have demonstrated the capabilities of a novel low-voltage electron microscope (LVEM) for imaging polymer and organic molecular thin films. The LVEM can operate in transmission electron microscopy, scanning transmission electron microscopy, scanning electron microscopy, and electron diffraction modes. The microscope operates at a nominal accelerating voltage of 5 kV and fits on a tabletop. A detailed discussion of the electron-sample interaction processes is presented, and the mean free path for total electron scattering was calculated to be 15 nm for organic samples at 5 kV. The total end point dose for the destruction of crystallinity at 5 kV was estimated at 5 x 10(-4) and 3.5 x 10(-2) C/cm2 for polyethylene and pentacene, respectively. These values are significantly lower than those measured at voltages greater than 100 kV. A defocus series of colloidal gold particles allowed us to estimate the experimental contrast transfer function of the microscope. Images taken of several organic materials have shown high contrast for low atomic number elements and a resolution of 2.5 nm. The materials studied here include thin films of the organic semiconductor pentacene, triblock copolymer films, single-molecule dendrimers, electrospun polymer fibers and gold nanoparticles.

Electrons↗
PubMed1985

Quaternary structure of pyruvate dehydrogenase complex from Escherichia coli.

The pyruvate dehydrogenase complex of Escherichia coli and subcomplexes derived from it by selective removal of component enzymes have been subjected to quaternary structural analysis by scanning transmission electron microscopy. Scanning transmission electron microscopic images of the intact complex (E1E2E3), the dihydrolipoyl transacetylase-dihydrolipoyl dehydrogenase (E2E3) subcomplex, and the E2 core enzyme appear as cubic particles in various orientations. Mass distributions within this complex and its subcomplexes have been determined by radial mass analysis of similarly oriented scanning transmission electron microscopic images of each type. The data show that mass attributable to dihydrolipoyl dehydrogenase (E3) is well integrated into the structural framework of the E2 core, dihydrolipoyl transacetylase, whereas mass attributable to pyruvate dehydrogenase (E1) is located about the periphery of the core enzyme. The mass distributions are fully consistent with a structural model in which 6 E3 dimers are integrated into the six faces of the cubic E2 core, and 12 E1 dimers are associated along the 12 edges of the core enzyme.

Escherichia coli↗
PubMed1983

Acute toxicity of lead particulates on pulmonary alveolar macrophages. Ultrastructural and microanalytical studies.

Although it is well established that respiratory uptake of lead-containing particles plays a substantial role in the epidemiology of plumbism, relatively little is known about the role of the pulmonary alveolar macrophage in lead poisoning. An in vitro system was designed to investigate the effects of lead oxide particles of respirable size on the rabbit alveolar macrophage. The studies were concerned with the intracellular solubility of PbO and Pb3O4 and changes in fine structure attributable to lead toxicity. The distribution of phagocytosed lead and its intracellular reprecipitation complexes was established by electron microprobe analysis and secondary ion mass spectroscopy in conjunction with transmission electron microscopy, scanning electron microscopy, scanning transmission electron microscopy, and backscatter imaging. It was found that Pb3O4, PbO and PbO-coated particles were ingested by the rabbit alveolar macrophages and that each of these lead oxide compounds produced similar damage to the fine structure of the cell. Swelling of the mitochondria, nuclear membrane, and endoplasmic reticulum was common, as well as were characteristic reprecipitation complexes of lead, phosphorous, and calcium within the nuclear heterochromatin and cytoplasm of the cell. The precipitation complexes were not seen in cells incubated with the particles if phagocytosis was blocked by 0.22-microns, membrane filters. It was concluded that phagocytosis of these lead oxide particles was necessary to produce the cytopathic changes. It is suggested that solubilization of lead from the ingested particles in phagosomes of macrophages results in the liberation of intracellular lead with the resultant formation of reprecipitation complexes.

Animals↗
PubMed1995

At least one of the protofilaments in flagellar microtubules is not composed of tubulin.

BACKGROUND: The core of the eukaryotic flagellum is the axoneme, a complex motile organelle composed of approximately 200 different polypeptides. The most prominent components of the axoneme are the central pair and nine outer doublet microtubules. Each doublet microtubule contains an A and a B tubule; these are composed, respectively, of 13 and 10-11 protofilaments, all of which are thought to be made of tubulin. The mechanisms that control the assembly of the doublet microtubules and establish the periodic spacings of associated proteins, such as dynein arms and radial spokes, are unknown. Tektins, a set of microtubule-associated proteins, are present in the axoneme as stable filaments that remain after the extraction of doublet microtubules; they are localized near to where the B tubule attaches to the A tubule and near to the binding sites for radial spokes, inner dynein arms and nexin links. Tektin filaments may contribute in an interesting way to the structural properties of axonemes. RESULTS: We have fractionated doublet microtubules from sea urchin sperm flagella into ribbons of stable protofilaments, which can be shown to originate from the A tubule. Using cryo-electron microscopy, conventional electron microscopy, scanning transmission electron microscopy, three-dimensional reconstruction and kinesin decoration, we have found that one protofilament in the ribbon is not composed of tubulin. This protofilament is an integral protofilament of the A tubule wall, has less mass per unit length than tubulin and does not bind kinesin. CONCLUSION: Contrary to what is generally assumed, at least one protofilament in the wall of the A tubule is not composed of tubulin. Our data suggest that this nontubulin protofilament is primarily composed of tektins, proteins that show some structural similarity to intermediate filament proteins. A 480 A axial periodicity within these ribbons, revealed by scanning transmission electron microscopy, can be related to the structure of tektin, and may determine the large-scale structure of the axoneme in terms of the binding of dynein, nexin and radial spokes to the doublet microtubule.

Animals↗
PubMed1985

A study of endometrial adenocarcinoma treated with tamoxifen by scanning and transmission electron microscopy.

By scanning and transmission electron microscopy the Authors studied four cases of endometrial adenocarcinoma (stage I, G1) after 15-days treatment with Tamoxifen (20 mg X 2) before surgery. The ultrastructural findings, similar to those observed in untreated adenocarcinomas but quite different from those obtained in MAP-responsive cases - as other Authors reported too - seem to indicate an almost complete absence of secretory or cytotoxic induction at least as far as 15-days treatment is concerned. According to the Authors this study raises many doubts about the usefulness of a first-instance therapeutical protocol based on Tamoxifen alone. However they believe that Tamoxifen can be utilized combined with a progestational agent in a simultaneous or sequence treatment.

Adenocarcinoma↗