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PubMed1981

The use of markers in immune electron microscopy.

Immune electron microscopy (IEM) cannot be used successfully for structures that do not have recognisable morphology. However, at least some of these structures or components are related antigenically to recognisable antigens or viruses. We have therefore mixed unknown antigens with known markers and looked for the presence of mixed aggregates. The present study examined a low molecular weight subunit of rotavirus and a micellar form of hepatitis B surface antigen. In both cases mixed immune aggregates were found showing that the unknown components had antigens in common with the established virus or antigen.

Antigen-Antibody Complex↗
PubMed1992

Has negative staining still a place in biomacromolecular electron microscopy?

Transmission electron microscopy of proteins has provided molecular- and in a few cases near-atomic-resolution structural information. In this review, we critically evaluate the potential and the limitations in obtaining molecular resolution, particularly with negatively stained specimens, and put these into perspective with cryomicroscopy of unstained frozen-hydrated and sugar-embedded preparations.

Histocytological Preparation Techniques↗
PubMed2005

Sequestering of p53 into DNA-protein filaments revealed by electron microscopy.

Using electron microscopy, we analyzed the interaction of bacterially expressed full-length p53, p53(1-393), and its C-terminal fragment, p53(320-393), with long (approximately 3000 bp) dsDNA in linear and supercoiled (|DeltaLk| approximately 4-6) forms containing or lacking the p53 recognition sequence (p53CON). The main structural feature of the complexes formed by either protein was a DNA-protein filament, in which two DNA duplexes are linked (synapsed) via bound protein tetramers. The efficiency of the synapse, reflected in its length and the fraction of molecules exhibiting DNA-protein filaments, was significantly modulated by the molecular form of the protein and the topological state of the DNA. With linear DNA, the synapse yield promoted by the C-terminus fragment was very low, but the full-length protein was effective in linking noncontiguous duplexes, leading to the formation of intramolecular loops constrained at their bases by short regions of synapsed DNA duplexes. When the linear DNA contained p53CON, regions of preferential sequence, i.e., encompassing p53CON and probably p53CON-like sequences, were predominantly synapsed, indicating a sequence specificity of the p53 core domain. With scDNA, the synapse yield was significantly higher compared to the linear counterparts and was weakly dependent on the sign of superhelicity and presence or absence of p53CON. However, the full-length protein was more effective in promoting DNA synapses compared to the C-terminal fragment. The overall structure of the DNA-protein filaments was apparently similar for either protein form, although the apparent width differed slightly (approximately 7-9 nm and approximately 10-12 nm for p53(320-393) and p53(1-393), respectively). No distortion of the DNA helices involved in the synapse was found. We conclude that the structural similarity of DNA-protein filaments observed for both proteins is attributable mainly to the C-terminus, and that the yield is dictated by the specific and possibly nonspecific interactions of the core domain in combination with DNA topology. Possible implications for the sequestering of p53 in DNA-protein filaments are discussed.

DNA↗
PubMed2005

Vertebral cancellous bone turn-over: microcallus and bridges in backscatter electron microscopy.

Backscatter electron microscopy (BSE) is a powerful technique for investigating cancellous bone structure. Its main function is to offer information regarding the degree of mineralization of the tissue within individual trabeculae. To illustrate the qualitative information that can be drawn from BSE imaging technique, we present a study on human vertebral cancellous bone. This tissue is continuously remodeled through osteoclastic resorption and osteoblastic new bone apposition. It is thought that osteoclastic resorption pits are especially deleterious for vertebral bone architecture since they often perforate the thin trabeculae; the osteoblasts being unable to repair the gap. In addition, excessive stress may also disrupt the architecture in case of trabecular fracture or damage accumulation. Waves of new bone formation were easy to identify in BSE. Often these waves were connecting both edges of a perforation and called bridges. Additionally, we present a few images of microcallus formations. A microcallus is described as a small mass of woven bone that generally repairs a trabecula. The microstructural aspects of different microcalluses are presented and discussed. Both bridges and microcallus should be considered as examples of the repair porcess since they obviously preserve the connectivity of the trabeculae. However, bridges were much more frequent than microcallus (396 vs 15). Both mechanisms probably illustrate the normal response to different local stimuli.

Adult↗
PubMedDate not supplied

Backscattered electron imaging: A new method for the study of cardiomyocyte architecture using scanning electron microscopy.

Scanning electron microscopy (SEM) with secondary electron emissions is useful for the study of cardiomyocyte architecture, however, the information is limited from the cell surface. Whereas backscattered electron (BSE) emission can give a high-resolution image of the specimen's intracellular structure after heavy metal staining. In this study, we applied BSE imaging analysis to the study of the arrangement of cardiomyocytes in the myocardium. The tissue specimens from a normal fresh monkey heart, normal human heart obtained at autopsy, and surgically resected tissue from a patient with old myocardial infarction in the left ventricular aneurysmectomy were used. The tissue specimens were fixed in neutral formalin, treated with NaOH and then stained with Gomori's silver methenamine reagent followed by tannic acid and osmium tetroxide. After dehydration and drying, the specimens were coated with carbon and examined by SEM with a BSE detector. In the tissue preparations, the A bands of sarcomeres were selectively stained with silver so that the arrangements of subsarcolemmal myofibrils and the intercalated discs were clearly seen in the BSE images. In the left ventricular aneurysmal walls of old myocardial infarction, atrophied cardiomyocytes with disarray of subsarcolemmal myofibrils were observed. The results strongly suggest that BSE images are further applicable to the study of the architecture of cardiac myocytes and their branches, and the arrangement of intracellular myofibrils in various diseased myocardium.

Animals↗
PubMed2000

Probing the building blocks of eumelanins using scanning electron microscopy.

Scanning electron microscopy (SEM) is used to examine the structure of natural and synthetic melanins. Eumelanin from Sepia officinalis and synthetic eumelanin are found to be structurally dissimilar. The natural sample has a significant structural order with subunits that have a lateral dimension of approximately 15 nm. The synthetic samples, on the other hand, appear to be amorphous solids. These results lend support for the existence of fundamental structural units proposed from the analyses of wide-angle X-ray diffraction measurements and previous mass-spectrometry results. These findings also provide insight into the disparate photophysical behavior of Sepia and synthetic eumelanin.

Animals↗
PubMed1994

Differential cell surface distribution of adhesion molecules demonstrated by immuno-scanning electron microscopy.

Scanning electron microscopy with immuno-gold labeling revealed that epidermal keratinocytes expressed ICAM-1 and HLA-DR molecules on their surfaces in patterns that differed in mycosis fungoides (MF) and lichenoid reaction (LR). ICAM-1 molecules, visualized as deposits of gold particles, were present as clusters adjacent to the junctions interconnecting the keratinocytes of MF lesions. LFA-1 molecules were seen as granules on the surfaces of all infiltrates, most of which also expressed ICAM-1. HLA-DR molecules were seen continuously along the borders of the individual keratinocytes. In LR, ICAM-1 and HLA-DR were expressed only sparsely on the undersurface of the epidermis, whereas the infiltrates expressed LFA-1 molecules on their surfaces. These findings may explain the differing histological features of MF and LR: ICAM-1 molecules present on the intercellular junctions of MF epidermis lead the LFA-1-bearing cells to migrate into the interspaces of the keratinocytes, thus producing "epidermotropism". These cells aggregate by means of co-expressed ICAM-1 to thus produce the appearance of a "microabscess". In LR, on the other hand, the minimal expression of ICAM-1 on the epidermal undersurface leaves most infiltrates within the dermis, thus producing a "band-like infiltrate" beneath the epidermis.

Aged↗
PubMed1979

The preservation of surface-associated micro-organisms prepared for scanning electron microscopy.

Scanning electron microscopy (SEM) has become a popular means of studying micro-organisms which associate with surfaces. However, as yet no detailed examination has been made of the influence of specimen preparation on the number of organisms finally seen on the SEM screen. In this investigation critical assessment is made of the influence of a wide range or preparative factors on the preservation of filamentous bacteria associated with the epithelial surfaces of rat intestine. Organisms were quantitated using a rigorous counting method (transect line analysis); statistical testing of these counts enabled the comparison of different preparative factors. The composition of the fixative was found to significantly influence the number of organisms preserved; of the fifteen fixatives studied, Karnovsky's fixative with ruthenium red best preserved surface-associated organisms. The influence of other factors on the number and appearance of preserved organisms was also examined. These factors included the washing of specimens prior to fixation, the storage of fixed specimens, and the handling and storage of critical point dried specimens. The results are discussed with reference to the optimal methods for preparing specimens for SEM.

Animals↗
PubMed1997

Effect of Escherichia coli L-form cytoplasmic membranes on the interaction between macrophages and Lewis lung carcinoma cells: scanning electron microscopy.

Scanning electron microscopy (SEM) investigations on the interactions between peritoneal macrophages from Lewis lung carcinoma (LLC)-bearing mice and LLC tumour cells during 21 days after tumour implantation were carried out. The action of lipopolysaccharide (LPS)-containing cytoplasmic membranes (CM), from the stable protoplast type L-form of Escherichia coli, on the activity of in vitro phagocytosis was studied; CM induced a continuous increase in macrophage numbers. Activation of macrophage surfaces in healthy and tumour-bearing mice was established. Lamelipods, pseudopods and migration fringes 14 days after CM application were seen. Crater-like cavities deeply in the macrophage cells as well as adherent or prominent engulfed tumour cells within macrophages were observed during in vitro interaction with LLC cells. Macrophages from tumour-bearing mice without CM treatment showed less activation evaluated by SEM during earlier stages of tumour growth. The SEM investigation proved the temporary stimulating effect of E. coli L-form CM on the cell surface activation of peritoneal macrophages in healthy and LLC-bearing mice.

Animals↗
PubMed2004

The benefits of using low accelerating voltage to assess endodontic instruments by scanning electron microscopy.

Scanning electron microscopy is often used to evaluate surface contamination and machining defects in dental and other medical instruments. Knowledge of the operating conditions of the SEM, in particular the accelerating voltage, is essential to properly interpret images of such material. We demonstrate the importance of using low accelerating voltages to detect surface features including contamination on NiTi rotary and hand files, and conclude that even recent studies may have significantly underestimated the amount of non-metallic debris (from the manufacturing process or from biological contamination) present on the surface of such instruments.

Dental Alloys↗
PubMed1974

Surface-surface associations in microbial communities populating epithelial habitats in the murine gastrointestinal ecosystem: scanning electron microscopy.

Scanning electron microscopy has been used to visualize the residents of microbial communities populating habitats on epithelial surfaces in the gastrointestinal tracts of mice. In the stomach, bacteria form a dense layer on the stratified squamous epithelium of the nonsecreting area. Microbes of at least three morphological types can be seen in this layer, including short rods with round ends, rods in chains, and tapering filaments composed of repeating units of rod- or coccal-shaped elements varying in size from large at one end of the filament to small at the other end. These three forms all attach by one end to the epithelium. The latter two forms can be found only so attached; in both cases, the end is inserted into a hole or depression in the keratinized epithelium. In the small intestine, a microbe of morphology similar to that of the tapering filaments found in the stomach can be seen attached end-on to the epithelium. Again each filament has one end inserted into a hole in the epithelium. In this case, however, the repeating elements of each filament are all about the same size. In the cecum and colon, predominantly fusiform- and spiral-shaped microbes can be seen mixed together in layers on the epithelium. At least three types of fusiform-shaped microbes can be distinguished on the basis of surface texture, and one type of spiral-shaped microbe can be found. These microorganisms appear to be attached to each other and to the epithelium by weblike filaments. The numerous microbial types present in the various epithelial habitats associate intimately surface-to-surface with each other and with the epithelium. Such surface-surface association may be an important autogenic factor contributing to the stability of the murine gastrointestinal ecosystem.

Animals↗
PubMed1973

Life cycle of Neurospora crassa viewed by scanning electron microscopy.

Scanning electron microscopy was used to examine the major stages of the life cycle of two wild-type strains of Neurospora crassa Shear and Dodge (St. Lawrence 3.1a and 74A): mycelia, protoperithecium formation, perithecia, ascospores, ascospore germination and outgrowth, macro and microconidia, and germination and outgrowth of macroconidia. Structures seen at the limit of resolution of bright-field and phase-contrast microscopes, e.g., the ribbed surface of ascospores, are well resolved. New details of conidial development and surface structure are revealed. There appears to be only one distinguishable morphological difference between the two strains. The pattern of germination and outgrowth which seems relatively constant for strain 74A or strain 3.1a, appears to be different for each. Conidia from strain 3.1a almost always germinate from a site between interconidial attachment points; whereas the germ tubes of strain 74A usually emerge from or very near the interconidial attachment site. These germination patterns usually do not segregate 2:2 in asci dissected in order. This observation suggests that conidial germination pattern is not under the control of a single gene.

Cell Differentiation↗
PubMed1974

Liberation and development of Allomyces arbuscula mitospores viewed by scanning electron microscopy.

Scanning electron microscopy has been employed to examine events in the release and development of mitospores of the aquatic fungus, Allomyces arbuscula. Among the salient features of spore release from the mitosporangium is the digestion of the inner matrix of the exit papillum. Hydrolysis appears to begin at the outer layer of the papillum plug matrix and probably results from activation of localized hydrolytic enzymes. The plug clearly consists of at least two different component layers. Elaboration of mitospores from the mitosporangium is depicted in several micrographs. Motile spores were induced to begin development, and the sequence of surface changes associated with the encystment process was studied. Time course studies show the retraction of the flagellum, the change from elipsoidal to spherical shape, and the deposition of the cell wall. Early in encystment, small vesicles accumulate on the surface of the plasma membrane. These enlarge and fuse to form the mature cyst wall. This surface view of cell wall deposition appears to support the possible role of gamma particles in cell wall synthesis during encystment.

Cell Membrane↗
PubMed1987

Topography, architecture and structure of the plexus submucosus internus (Meissner) of the porcine small intestine in scanning electron microscopy.

Scanning electron microscopy of whole-mount preparations of the tela submucosa in the porcine small intestine, examined after trypsin digestion, fixation and HCl hydrolysis, visualized a clear differentiation of the submucosal plexuses, i.e., the plexus submucosus internus (Meissner) and the plexus submucosus externus (Schabadasch). The distinctive features refer to the topography, number, size and shape of the ganglia and the number and diameter of the nerve strands. The plexus of Meissner is closely apposed to the external surface of the lamina muscularis mucosae by the enveloping connective tissue and by connecting strands penetrating the lamina muscularis mucosae. Three distinctive subdivisions of connecting strands can be identified. Since the glial cells covering the ganglia and connecting strands have been preserved, neither individual neuronal cells nor axons can be observed.

Animals↗
PubMed1986

The microvascular localization of laminin and fibronectin in scleroderma muscle by immune electron microscopy.

Immune electron microscopy using a peroxidase-antiperoxidase (PAP) technique was done on muscle biopsies from patients with scleroderma (PSS), rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), and normal controls, in order to localize laminin and fibronectin in the muscle capillary basement membrane (BM). Three patterns of staining were noted for the 2 antigens: in/out, diffuse and negative. Staining patterns in PSS biopsies were markedly different from both the normal and disease controls. For laminin the expected in/out pattern was rare in PSS, with virtually all biopsies being either diffuse or negative. In the case of fibronectin, the staining patterns were of equal frequency in PSS, whereas in the control groups diffuse staining was rare. These findings suggest that in addition to the morphologic abnormalities previously noted in PSS muscle capillary BM, there is also an alteration in the distribution of specific BM components in PSS.

Arthritis, Rheumatoid↗
PubMed2002

Evidence for Escherichia coli O157:H7 attachment to water distribution pipe materials by scanning electron microscopy.

Scanning electron microscopy observation was used to investigate the adhesion of Escherichia coli O157:H7 on water distribution pipe surfaces such as copper and polyethylene plastic at different contact times and storage temperatures. Our results indicated that E. coli cells could easily attach to both surface types after exposures as short as 1 or 4 h at ambient (20 degrees C) and refrigeration temperatures (4 degrees C). Also, we found that copper surfaces have a higher number of attached E. coli cells than plastic surfaces. The number of cells attached to each type of material depended on the nature of the water distribution pipe surfaces and the length of contact time. In addition, the surface energy value of each surface estimated by contact angle measurements using water, alpha-bromonaphthalene, and dimethyl sulfoxide as wetting agents showed that both copper (41.2 megajoules [MJ] m(-2)) and plastic (45.8 MJ x m(-2)) have a low energy surface. In no cases could evidence of extracellular material be observed on surfaces with either exposure condition.

Bacterial Adhesion↗
PubMed1999

[LASIK--comparison of oscillating and rotating microkeratotomy using scanning electron microscopy].

Scanning electron microscopy was used to evaluate the differences in the quality of the cut produced by oscilating microkeratome Flapmaker and Draeger rotating microkeratome. Higher quality of the edge of the cut was found with Flapmaker. Both microkeratomes produce uniform surface of the cut. Chatter is much less expressed with Flapmaker. No difference at the interface tissue was found with SEM three weeks postoperatively comparing the microkeratomes.

Animals↗