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PubMed1990

Computerized three-dimensional reconstructions of serial sections in electron microscopy.

To improve reconstructive 3D electron microscopy novel methods are discussed to represent and process serial section images in a cuberille environment. This includes the analysis of the transfer characteristics of the image detection system, the use of laser-induced fiducials for deformation correction and alignment, the control of section thickness by EELS and the use of ESI to image thick sections.

Algorithms↗
PubMed1982

Scanning electron microscopy.

The technology of scanning electron microscopy (SEM) is described in brief. Its application to the study of cell and tissue structure is demonstrated and the evolution of the concept of "topographical histology" is discussed. Some current literature on the application of SEM is reviewed under the headings of experimental pathology and human pathology. While SEM has become an indispensable technique for the experimental morphologist, its application to diagnostic pathology and cytology is still at an early exploratory stage.

Animals↗
PubMed1994

Study on gutta-percha points using scanning electron microscopy and analysis with electron microprobe.

A comparative study of the apical morphology and contour was performed with the scanning electron microscope at 45x, 70x and 300x, while the chemical composition of eight gutta-percha point brands was surveyed with an EDAX 9100 electron microprobe. They were classified according to their apical morphology as conical (25%) and truncate (75%), based on their contour as regular (50%) and irregular (50%) and according to their surface as without defects (50%) and with defects (50%). The analysis of inorganic compounds revealed the presence of varying proportions of Zn, Ba, Si, Mg. Ca, P, Cl and Al, possibly implying that the presence of Ba and Al could interfere with postendodontic repair or at least irritate the periapical area if inadvertently overfilled.

Electron Probe Microanalysis↗
PubMed1988

Mapping gold-labeled IgE receptors on mast cells by scanning electron microscopy: receptor distributions revealed by silver enhancement, backscattered electron imaging, and digital image analysis.

Immunogold labeling and silver enhancement techniques are widely used to determine density and distribution of cell membrane receptors by light and transmission electron microscopy. However, these techniques have not been widely used for receptor detection by scanning electron microscopy. We used antigen- or protein A-conjugated colloidal gold particles, together with silver enhancement, sequential secondary and back-scattered electron imaging (SEI and BEI), and digital image processing, to explore cell surface distribution of IgE-receptor complexes on RBL-2H3 cells, a rat leukemia line that provides a model for the study of mucosal mast cells. Cells were first incubated with a monoclonal antidinitrophenol IgE (anti-DNP-IgE) that binds with high affinity to cell surface IgE receptors. The resulting IgE-receptor complexes were cross-linked either with the multivalent antigen, DNP-BSA-gold, or with a polyclonal anti-IgE antibody. Antibody-treated cells were labeled after fixation with protein A-gold. Fixed, gold-labeled cell monolayers were silver enhanced (or not), dehydrated, critical point-dried, and coated with gold-palladium (for SEI analysis) or carbon (for combined SEI/BEI analysis). They were observed in an Hitachi S800 SEM equipped with a field emission tip and a Robinson backscattered electron detector. An image processor (MegaVision 1024XM) digitized images directly from the S800 microscope at 500-1000 line resolution. Silver enhancement significantly improves detection of gold particles in both SEI and BEI modes of SEM. On gold-palladium-coated samples, 20-nm particles are resolved by SEI after enhancement. BEI resolves 15-nm particles without enhancement and 5- or 10-nm particles are resolved by BEI on silver-enhanced, carbon-coated samples. Neither BEI nor SEI alone can yield high resolution topographical maps of receptor distribution (BEI forms images on the basis of atomic number contrast which reveals gold but not surface features). Image analysis techniques were therefore introduced to digitize, enhance, and process BEI and SEI images of the same field of view. The resulting high-contrast, high-resolution images were superimposed, yielding well-resolved maps of the distribution of antigen-IgE-receptor complexes on the surface of RBL-2H3 mast cells. The maps are stored in digital form, as required for computer-based quantitative morphometric analyses. These techniques of silver enhancement, combined BEI/SEI imaging, and digital image analysis can be applied to analyze density and distribution of any gold-labeled ligand on its target cell.

Cell Line↗
PubMed1996

Diagnostic value of electron microscopy in rare malignant musculoskeletal tumors. Experience from an orthopedic tumor center.

Rare musculoskeletal tumors can be difficult to diagnose by light microscopy or immunohistochemistry. Electron microscopy can be of diagnostic assistance especially if histotype specific ultrastructural features exist. In particular, electron microscopy for uncommon sarcomas such as alveolar soft part sarcoma, parachordoma, atypical Ewing's sarcoma and epithelioid sarcoma may be the diagnostic modality of choice.

Adult↗
PubMed1985

The ultrastructure of the electric burn in man: a transmission electron microscopy-scanning electron microscopy study.

The authors describe the ultrastructural aspects of the electric burn observed with the transmission electron microscope (TEM) and the scanning electron microscope (SEM). With the TEM the most apparent feature is the homogenization and arrangement of the filamentous cytoplasmic material in elongated and parallel bundles together with elongation of the nuclei and junctional structures. SEM studies present a three-dimensional vision of the filamentous material that confirms and clarifies the cellular deformation as a whole. An hypothesis is advanced suggesting that the phenomenon is due to the effects produced by the extremely rapid dehydration of the tissue or that the electric current and Joule effect can act upon the filaments and contractile proteins, causing the cell deformation.

Burns, Electric↗
PubMed1975

Education and training in electron microscopy.

There is currently a need for diagnostic electron microscopy in both autopsy and surgical pathology. As more information emerges from the research laboratories, applied electron microscopy will grow in depth, and all large medical centers will need the expertise provided by a diagnostic electron microscopy laboratory. Many physicians other than electron microscopists find it essential to have an understanding of the contributions and limitations of electron microscopy. Education and training programs therefore must encompass not only paramedical personnel who prepare the electron micrographs and pathologists who are thoroughly trained in interpret the electron micrographs, but also other physicians and scientists who utilize the information obtained therefrom. Medical students should obtain sufficient background in normal and abnormal ultrastructure to enable them to interpret the medical literature, and the means to obtain this background should be available within the medical school curriculum. Some medical students will also desire more thorough training obtained by elective courses in electron microscopy. Pathology residents should obtain sufficient expertise during their residency to enable them to utilize the information produced by the electron microscopy laboratory. Certain pathology residents and some pathologists in practice prefer fellowships for more specialized training in electron microscopy. Four representative training programs in electron microscopy from Veterans Administration hospitals have been selected for presentation. Each emphasizes a different approach and different objectives.

District of Columbia↗
PubMed1993

Localization of DNA topoisomerase II in Chinese hamster fibroblasts by confocal and electron microscopy.

The localization of the 170- and the 180-kDa isoforms of the enzyme DNA topoisomerase II in growing Chinese hamster fibroblasts has been studied by confocal immunofluorescence microscopy and immunogold electron microscopy after labeling with affinity-purified isoform-specific polyclonal antibodies. Immunofluorescence and immunogold studies, together with quantitative image analysis, show that the two isoforms are present in the nucleoplasm and in the nucleolus. In the nucleoplasm both isoforms are frequently localized at the periphery of heterochromatin regions. In the nucleolus the immunofluorescence and immunogold signals relative to surface area are higher than in the nucleoplasm; both isoforms are localized predominantly in the fibrillar zones. During mitosis both isoforms remain detectable in the cytoplasm. The differential expression of the two isoforms during the cell cycle, observed in other studies, suggests that they have different functions, and their presence in both the nucleoplasm and the nucleolus suggests that these functions are required in both of these nuclear compartments.

Animals↗
PubMedDate not supplied

Detection of VP-16-treated HL-60 cell apoptosis by TUNEL electron microscopy.

Apoptosis is essential to many physiological processes, including maturation of cells and the immune system. Deficient regulation of apoptosis may play an important role in many pathological conditions, such as autoimmunity, AIDS, and myelodysplastic syndrome. Several methods have been described to identify apoptotic cells. DNA strand breaks can be identified by labeling free 3'-OH termini with modified nucleotides by enzymatic reaction (TUNEL method). In this study, apoptosis was introduced in cultured HL-60 cells treated with VP-16, and the TUNEL method was adapted for electromicroscopy, called the EM TUNEL method. The results of the EM TUNEL method were compared with those of light microscopy and flow cytometry. Apoptotic cells following VP-16 (10 micrograms/mL) treatment were detected after 3 h by all methods (light microscopy, Erythrocin B, electron microscopy, flow cytometry, TUNEL, and EM TUNEL). EM TUNEL was the most sensitive method of detection, with a detection rate of 32%. Furthermore, EM TUNEL was more suitable for distinguishing cell lineage than light microscopy TUNEL. The results indicate that EM TUNEL can be used to detect apoptosis in bone marrow species in vivo.

Apoptosis↗
PubMed1997

A reevaluation of routine electron microscopy in the examination of native renal biopsies.

Electron microscopy is routinely utilized in most centers in the evaluation of native renal biopsies. Several studies, primarily from the 1960s and early 1970s, provide justification for its use. Conducted by Siegel et al. (1), the largest study evaluated 213 consecutive renal biopsies and found that electron microscopy was needed for a correct diagnosis in 11%, as well as for confirmation or additional information in another 36%. However, nearly all of these studies were conducted before the use of immunofluorescence in renal biopsy diagnosis became widespread and before several new glomerular diseases and variants were described. In light of this situation and the expense of the procedure, the routine use of electron microscopy in native renal biopsies also examined by immunofluorescence and routine light microscopy was reevaluated. From January 1996 to June 1996, 288 native renal biopsies were received, and all were evaluated by the same pathologist. Of those, 233 met criteria for inclusion in this study, which were > or = 5 glomeruli for light microscopy, > or = 2 for immunofluorescence, and > or = 1 for electron microscopy, not including globally scarred glomeruli. Light microscopy (hematoxylin and eosin, periodic acid-Schiff stains) and immunofluorescence--for immunoglobulin (Ig) G, IgA, IgM, C3, C1q, fibrinogen; kappa/lambda when needed--were evaluated on each biopsy within 48 h of receipt, and a preliminary diagnosis was recorded if possible. Electron microscopy was then performed, and a final diagnosis was made. In 50 cases (21%), electron microscopy was needed to make the final diagnosis; in two of these cases, the preliminary diagnosis was incorrect, and in 48, a firm preliminary diagnosis could not be made. In the other cases, the preliminary diagnosis was correct, but in 48 (21%), ultrastructural study was felt to provide important confirmatory data, and in eight cases (3%), an additional, unrelated diagnosis was supported by the ultrastructural findings. Diagnoses most frequently requiring electron microscopy included minimal change nephropathy, early diabetic nephropathy, membranous lupus nephritis, membranoproliferative glomerulonephritis, postinfectious glomerulonephritis, thin basement membrane nephropathy (or exclusion of this in cases of otherwise unexplained hematuria), and human immunodeficiency virus-associated nephropathy (or exclusion of it in cases of collapsing glomerulopathy). Common diagnoses usually not requiring electron microscopy included IgA nephropathy, diffuse proliferative lupus nephritis, focal segmental glomerulosclerosis (not collapsing glomerulopathy variant), pauci-immune crescentic glomerulonephritis, acute interstitial nephritis, and amyloid nephropathy. This study confirms that, as was the case 20 to 30 yr ago, electron microscopy provides useful diagnostic information in nearly half of native renal biopsies. If electron microscopy cannot be performed routinely on all such biopsies, it is recommended that tissue for ultrastructural studies be set aside in each case.

Biopsy↗
PubMed1998

[Fetal hypotrophy: morphological pictures of placenta in light and electron microscopy of female residents of the Czenstochow region].

The 16 afterbirths samples obtained from pregnancies complicated by idiopathic fetal hypotrophy was examined. The criteria of hypotrophy were established: the duration of gestation above 36 weeks, the birth-weight was less than 2500 g. The control group included 8 afterbirth samples from physiological full-term pregnancies. It was performed macroscopic examination, light microscopy and transmission electron microscopy. Macroscopically, no changes were found in the study group apart from the small differences of the weight. Light microscopy showed the retardation of placenta maturation from fetus hypotrophy group, congestion of the villi and presence of the numerous calcifications. Transmission electron microscopy showed the changes of placental barrier elements: adaptation processes and chronic progressive damage of the cells (exfoliation of the endothelial cells from the basal membrane).

Adult↗
PubMed1993

Lamin B distribution and association with peripheral chromatin revealed by optical sectioning and electron microscopy tomography.

We have used a combination of immunogold staining, optical sectioning light microscopy, intermediate voltage electron microscopy, and EM tomography to examine the distribution of lamin B over the nuclear envelope of CHO cells. Apparent inconsistencies between previously published light and electron microscopy studies of nuclear lamin staining were resolved. At light microscopy resolution, an apparent open fibrillar network is visualized. Colocalization of lamin B and nuclear pores demonstrates that these apparent fibrils, separated by roughly 0.5 micron, are anti-correlated with the surface distribution of nuclear pores; pore clusters lie between or adjacent to regions of heavy lamin B staining. Examination at higher, EM resolution reveals that this apparent lamin B network does not correspond to an actual network of widely spaced, discrete bundles of lamin filaments. Rather it reflects a quantitative variation in lamin staining over a roughly 0.5-micron size scale, superimposed on a more continuous but still complex distribution of lamin filaments, spatially heterogeneous on a 0.1-0.2-micron size scale. Interestingly, lamin B staining at this higher resolution is highly correlated to the underlying chromatin distribution. Heavy concentrations of lamin B directly "cap" the surface of envelope associated, large-scale chromatin domains.

Animals↗
PubMed1997

A leiomyoma of the iris documented by immunohistochemistry and electron microscopy.

PURPOSE: To report an iris leiomyoma documented in accordance with today's stricter diagnostic criteria. METHODS: Light microscopy, immunohistochemistry and electron microscopy were performed on the 3 mm ball-like, greyish-white vascularized tumour of the iris, close to the sphincter pupillae. RESULTS: Light microscopy of the extirpated neoplasm showed the characteristic appearance of a leiomyoma with densely packed spindle-shaped cells, with oval nuclei and granular cytoplasm. On electron microscopic examination the tumour exhibited the characteristic features of a smooth muscle neoplasm. The immunohistochemistry was consistent with a myogenic tumour because the tumour cells were positive for smooth muscle actin and desmin, but negative for S-100 and melanin. CONCLUSION: The case illustrated the necessity of performing ancillary procedures such as electron microscopy and immunohistochemistry to substantiate a correct, although rare diagnosis.

Actins↗
PubMed1993

Solution structure and direct imaging of fibronectin adsorption to solid surfaces by scanning force microscopy and cryo-electron microscopy.

In this study, we present the scanning force and electron microscopic visualization of single molecules of fibronectin either frozen hydrated or adsorbed onto metallic and polymeric surfaces with different solid surface tensions. The surfaces were characterized by dynamic contact angle measurements, X-ray photo emission spectroscopy (XPS or ESCA) and scanning force microscopy. The proteins were prepared by fast protein liquid chromatography (FPLC) and characterized by gel electrophoresis. Protein films on surfaces were investigated by surface plasmon resonance spectroscopy and directly imaged by scanning force microscopy. The spreading of the adsorbed fibronectin revealed dependence on the chemical composition and the solid surface tension. Structure of fibronectin in solution as well as on solid interface appeared as an extended straight strand as obtained by imaging with electron and scanning probe microscopies. Imaging of DNA was performed by scanning force microscopy to test the accuracy and reproducibility of our measurements. The measured contour lengths were accurate and the larger widths were caused by convolution of the tip shape and sample. Frictional forces during the scan have been of significant contribution in the imaging mechanism. Moreover, this work demonstrated that scanning force microscopy can be used for mapping the orientation and organization of protein film adsorbed onto various surfaces at the nanoscale.

Adsorption↗
PubMed2006

Noise-induced gastric lesions: a light and scanning electron microscopy study of the alterations of the rat gastric mucosa induced by low frequency noise.

INTRODUCTION: Exposure to low frequency noise (LFN) can lead to vibroacoustic disease (VAD), recognized as a systemic disease with lesions in a broad spectrum of organs. Although gastrointestinal complaints are common among individuals exposed to noise, only few studies tried to evaluate the digestive lesions. The authors performed this study in order to investigate gastric lesions in an animal model of VAD. MATERIAL AND METHODS: Adult Wistar rats were exposed to continuous LFN. After five weeks they were sacrificed. The stomachs were studied by light microscopy and scanning electron microscopy, and compared with stomachs of animals kept in silence. RESULTS: Superficial erosions were present in the noise-exposed animals. Massive cell death of the gastric epithelium was observed, both by light and electron microscopy. DISCUSSION: The erosions, reflecting cellular degeneration and death, occurred without inflammation, similar to what has been observed in other LFN-exposed organs.

Animals↗
PubMed1999

An effective method for suspicious cases in urinary cytodiagnosis by the combined use of scanning and transmission electron microscopy following light microscopy.

In urinary cytodiagnosis, pathological characterization of atypical cells is sometimes difficult because cells in urine samples tend to be degenerated. To overcome this problem, we adopted serial examination under light microscopy (LM), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). Five patients with transitional cell carcinoma primarily developed in the bladder, three patients with dysplastic cells, and one patient with metastatic and infiltrative colorectal carcinoma were subjects of the present study. Sample cells were smeared on a film slide especially suitable for serial use (a film slide used in X-ray diagnosis), fixed in 1% glutaraldehyde (GA) for 30 min, stained with Papanicolaou (Pap) staining, and immediately processed for fixation in 2% paraformal-dehyde (PA) and 2.5% GA. Slides were postfixed in 0.5% osmium tetroxide. Then, cell samples were subjected to a series of observations under LM, SEM, and TEM. One of three patients in whom dysplastic cells were suggested under LM was finally diagnosed as transitional cell carcinoma by subsequent examination under SEM followed by TEM. Thus, serial use of LM, SEM, and TEM proved useful in discriminating transitional cell carcinoma (grade 1) from dysplastic cells and also in distinguishing metastatic tumor cells from primary tumor cells in urine samples.

Journal Article↗
PubMed1997

Role of electron microscopy in interstitial lung disease.

Although electron microscopy no longer enjoys the important role in the diagnosis of interstitial lung diseases that it had in the 1960s and 1970s, it remains an important adjunct in the differential diagnosis of certain pulmonary diseases. Examples include various manifestations of systemic lupus erythematous pneumonitis, in which the presence of tubuloreticular structures and electron-dense deposits are useful for diagnosis; immotile cilia disorders, in which qualitative and now quantitative studies of the cilia of respiratory epithelial cells can help to establish the diagnosis; infections by viruses and other subcellular microorganisms as shown by the role played by electron microscopy in the initial diagnosis of the Hantavirus pulmonary syndrome; pneumoconioses, in which, in conjunction with elemental analysis probes, scanning electron microscopy is of critical importance in establishing the presence of offending foreign compounds in lung tissue or fluids; pulmonary fibrilloses, such as amyloidosis, light chain disease, and fibrillary glomerulonephritis, affecting the lung; and cases of alveolar proteinosis or Langerhans cell granulomatosis diagnosed from fluids such as bronchoalveolar lavages or small tissue samples. As important, electron microscopy remains of enormous usefulness in the study of early structural events leading to the pathogenesis of diseases. For example, recent uses of the technique have focused on the alveolar-capillary wall damage induced by alveolitis in hypersensitivity pneumonitis and sarcoidosis. In summary, electron microscopy remains a useful method in the study and diagnosis of some interstitial lung diseases, but because of its expense it is incumbent on the clinician to use good judgment in the selection of cases and diseases for study by this method.

Diagnosis, Differential↗