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At least 973 records · Page 54Linked to original sources
PubMed2005

Visualization of the nuclear lamina in mouse anterior pituitary cells and immunocytochemical detection of lamin A/C by quick-freeze freeze-substitution electron microscopy.

We examined the nuclear lamina in the quickly frozen anterior pituitary cells by electron microscopic techniques combined with freeze substitution, deep etching, and immunocytochemistry and compared it with that in the chemically fixed cells. By quick-freeze freeze-substitution electron microscopy, an electron-lucent layer, as thick as 20 nm, was revealed just inside the inner nuclear membrane, whereas in the conventionally glutaraldehyde-fixed cells the layer was not seen. By quick-freeze deep-etch electron microscopy, we could not distinguish definitively the layer corresponding to the nuclear lamina in either fresh unfixed or glutaraldehyde-fixed cells. Immunofluorescence microscopy showed that lamin A/C in the nucleus was detected in the acetone-fixed cells and briefly in paraformaldehyde-fixed cells but not in the cells with prolonged paraformaldehyde fixation. Nuclear localization of lamin A/C was revealed by immunogold electron microscopy also in the quickly frozen and freeze-substituted cells, but not in the paraformaldehyde-fixed cells. Lamin A/C was localized mainly in the peripheral nucleoplasm within 60 nm from the inner nuclear membrane, which corresponded to the nuclear lamina. These results suggest that the nuclear lamina can be preserved both ultrastructurally and immunocytochemically by quick-freezing fixation, rather than by conventional chemical fixation.

Animals↗
PubMed2002

Scanning near field optical/atomic force microscopy of bromodeoxyuridine-incorporated human chromosomes.

The present study applied scanning near field optical/atomic force microscopy (SNOM/AFM) to the observation of human chromosomes immunostained with an anti-BrdU antibody after incorporation of BrdU into DNA. Human lymphocytes were cultured in BrdU for 72 h and their chromosomes were prepared with a standard method for light microscopy. After additional fixation with 15% formalin in phosphate buffered saline, the specimens were denatured with 2N HCI with 0.1% Triton-X 100, immunostained with the anti-BrdU antibody, and observed both by fluorescence microscopy and by SNOM/AFM. The preparation technique used in the present study enabled the differential staining of sister chromatids in each chromosome, and sister chromatid exchanges (SCEs) were recognized in some chromosomes of the metaphase spread. Observations of the specimens by SNOM/AFM further provided the simultaneous collection of topographical and fluorescent images of the same portions of BrdU-incorporated chromosomes. The resolution of the fluorescence images by SNOM/AFM was greater than that obtained by fluorescence microscopy. Superimposition of topographical and fluorescent images of the chromosomes is useful for the precise analysis of the fine structure of chromosomes in relation to the SCEs. The application of SNOM/AFM to the BrdU-incorporated chromosomes is thus useful for the analysis of the fine structure of chromosomes in relation to their function.

Antimetabolites↗
PubMed1993

Correlative light and electron microscopy of the same sections embedded in HPMA, Quetol 523 and MMA.

Semithin sections, cut from tissues stained with acid and basic dyes after embedding in 2-hydroxypropyl methacrylate, Quetol 523 and methyl methacrylate, showed cytoplasmic components at a high resolution by light microscopy. These same sections could then be viewed, after osmium tetroxide, uranyl and lead staining, by the electron microscope. These sections had a number of inherent advantages: they could be observed with a light microscope; they facilitated analysis of cellular structures in the identical sites, and they were frequently the optimum thickness to provide three-dimensional information. We clearly established the structural detail of this same-section correlative light-electron microscopy approach by showing that the coloured materials observed in such sections of cells followed the distribution of fine structures within the same sections as determined by electron microscopy. In some instances the fidelity of the correlation between the distribution of the coloured area and cytoplasmic components in identical cells of the same section revealed significant details which could not visualized in thin sections. This technique, therefore, provided a simple and useful solution to many problems that require the localization of cellular components in identical cells selected previously by light microscopy.

Animals↗
PubMedDate not supplied

Correlative light optical, scanning electron, and transmission electron microscopy of skeletal muscle in muscular dystrophy and muscular atrophy: a pilot study.

Biopsies of skeletal muscle from three different cases of muscular dystrophy and one case of spinal muscular atrophy that has been fixed with Karnovsky's fluid were either routinely prepared for scanning electron microscopy (SEM) or were frozen to -20 degrees C and sectioned on a steel knife in a cryostat at 5-10 micrometer. The sections were coverslipped and examined using a light microscope equipped with polarizing optics (Pol). After areas were selected, the sections were prepared for SEM and thereby examined. The tissues on the slides that had been observed with light microscopy (LM) and SEM were prepared further for transmission electron microscopy (TEM) by infiltrating them with Epon and cutting sections at approximately 100 nm on an ultramicrotome. It is shown that the stage of contraction in one pathologic myofiber may vary along its length. The following advantages may be realized by using correlative (Pol leads to SEM leads to TEM) microscopy on skeletal muscle biopsies: 1) lesions can be differentiated from "normal" surrounding tissue; 2) doubtful structures can be reexamined with the SEM and TEM; and 3) the SEM image of different states of muscle contraction can be reinterpreted in the light of the Pol or TEM image.

Adolescent↗
PubMed2000

Use of a Technovit 7200 VLC to facilitate integrated determination of aluminum by light and electron microscopy.

Technovit 7200 VLC is an excellent embedding medium for both inorganic histochemistry by light microscopy and X-ray microanalysis by scanning and transmission electron microscopy. Liver samples from rats after intraperitoneal treatment with aluminum chloride were fixed in glutaraldehyde and embedded in the resin. Thick sections were easily cut on an ultramicrotome and stained with aluminon for aluminum (Al). An intense positive reaction with aluminon was observed in the Kupffer cells by light microscopy. The surface structures of the same resin block cut for light microscopy were observed under a scanning electron microscope fitted with an energy dispersive X-ray spectrometer. The Kupffer cells appeared white in the backscattered mode. Localization of Al in the Kupffer cells was confirmed by an X-ray distribution map in the scanning electron microscope. Subcellular localization of Al in the Kupffer cells was performed on the same semithin sections using a transmission electron microscope equipped with an energy dispersive X-ray spectrometer. Most Al was found in lysosomes of the Kupffer cells. The resin was stable in the electron beam and chlorine-free.

Acrylic Resins↗
PubMed1998

The value of fluorescence microscopy of auramine stained sputum smears for the diagnosis of pulmonary tuberculosis.

Laboratory diagnosis of pulmonary tuberculosis rests on the bacteriological examination of sputum smears stained by the Ziehl-Neelsen (ZN) method for acid fast bacilli (AFB). In the present study, we have compared light microscopy of ZN stained smears with that of fluorescence microscopy of sputum smears stained by auramine-phenol flurochrome dye for detection of AFB in sputum specimens. Sputum specimens from a total of 2,600 clinically suspected and diagnosed cases of pulmonary tuberculosis were examined by both the methods. Sputum specimens from a total of 1,104 patients were found to be positive for AFB. These included sputa from 975 (37.5%) patients positive for AFB by both ZN and auramine staining methods and sputa from an additional 129 (4.96%) patients positive for AFB by auramine staining only. Thus auramine staining of sputum smears in comparison to that of ZN staining is a better method of sputum microscopy for demonstration of AFB in sputum specimens. Fluorescence microscopy is relatively more sensitive and has the added advantage of allowing a large number of sputum specimens to be examined in a given time, in laboratories equipped with a fluorescent microscope.

Bacteriological Techniques↗
PubMed2000

Confocal and multi-photon microscopy of dental hard tissues and biomaterials.

Confocal microscopy is a technique that can be used both in the clinic and the high-resolution microscopy suite. This form of optical microscopy enables high-resolution images to be made of samples with minimum requirements for specimen preparation. Images may be made of either reflections from the sample surface or, if an immersion medium is used to optically couple the objective lens, then sub-surface images can be produced of reflective or fluorescent structures within semi transparent materials such as cells and dental hard tissues. These images are like optical sections, giving thin (> 0.35 microm) slices up to 200 microm below the surface of a mineralized tissue. The technique generates significant improvements in resolution, lying somewhere between that of conventional light microscopy and TEM/SEM. Instruments that work at video-rate allow high-speed events to be examined, such as in vivo clinical studies, cutting of dental tissues and fracture of adhesive interfaces. New dyes offer many exciting prospects for labeling changes in chemical composition in materials or biological tissues, while new imaging techniques such as multi-photon laser excitation of dyes give the potential of greater depth penetration and improved resolution. As with all new techniques the inexperienced should be aware of some of the artifacts inherent to the system. However, the widespread availability of conventional confocal microscopes should give ample opportunity for dental researchers to capitalize on this new technology.

Artifacts↗
PubMed2000

[Observation of biofilms inside tracheal tubes by electron microscopy and the relationship between biofilms and VAP].

OBJECTIVE: To observe the formation of biofilms inside tracheal tubes and to evaluate the effect of biofilm on the development of ventilator associated pneumonia (VAP). METHODS: Biofilms inside tracheal tubes from intubated patients were observed by scanning electron microscopy and transmission electron microscopy, meanwhile bacteria were detected in the specimens from the lower respiratory tract and the inside of tracheal tubes. RESULTS: 76% (19/25) of inner surface of tracheal tubes was coated with a confluent amorphous matrix by scanning electron microscopy. The average duration of trach-intubation in this group [(10.7 +/- 7.9) days] was longer than that in the other group [(2.1 +/- 0.8) days] in which tracheal tubes were not coated with the confluent amorphous matrix (P < 0.05). The presence of many bacteria(14/18) within these amorphous matrix was confirmed by transmission electron microscopy. Of the sixteen tracheal tubes, eleven tubes grew the same organisms that had been isolated from the secretions of the lower respiratory tract before extubation. In some VAP patients (7/8), organisms isolated from tracheal tubes were the same organisms which were pathogenic organisms of VAP. CONCLUSIONS: There was a close connection between the formation of bacterial BF and the long-term indwelling tracheal tubes. The presence of bacterial BF in the inner of tracheal tubes might be associated with the pathogenic organisms of VAP.

Adult↗
PubMed2001

Confocal laser scanning, conventional scanning and transmission electron microscopy of vertebrate cerebellar granule cells.

Confocal laser scanning microscopy of hamster cerebellar granular layer showed in montages of z-series the presence of small, medium and large granule cells. A granule cell Golgi cell ratio of 50/4 was observed surrounding glomerular regions. Field emission high resolution scanning electron microscopy of mouse cerebellar granular and molecular layers showed SE-I images of the outer and inner surfaces of nuclear and cytoplasmic compartments of chromium coated granule cells and the axo-spinodendritic synapses of parallel fibers with Purkinje cell dendrites. Conventional scanning electron microscopy of teleost fish cerebellar cortex showed three dimensional morphology of granule cell soma and processes and the synaptic relationship with mossy and climbing fibers, Golgi cell axonal ramifications and dendrites of stellate neurons, by means of SE-II and SE-III signal image mode, in sagittally and transversally cryofractured cerebellar cortex. SE-II images of the non-synaptic segments and synaptic varicosities of parallel fiber outer surface were characterized in the molecular layer. Ultrathin sections of transmission electron microscopy (TEM) revealed somato-somatic, dendro-somatic and dendro-dendritic like-desmosomal and like-hemidesmosomal junctions in human cerebellar granule cells. Freeze-etching replicas of mouse cerebellar cortex displayed granule cell intramembrane morphology, cytoplasmic fractured face and the Bergman glial cell cytoplasm completely surrounding the parallel fibers in the molecular layer. The mossy fiber-granule cell dendrite synaptic relationship was observed in sagittally and transversally cryofractured cerebellar cortex and correlated with TEM images. SE-II images of the climbing fiber synaptic connections with granule cell dendrites were obtained in teleost fish cerebellar cortex. One to one axo-dendritic synaptic contacts between Golgi cell axonal ramifications and granule cell dendrites were also seen. The above findings provide new vistas for future studies dealing with intracortical circuits and information processing in the cerebellar cortex.

Animals↗
PubMed2004

[A comparative study of malignant tissue diagnosis using ATR and microscopy FTIR spectroscopy].

A comparative FTIR study was undertaken between ATR (attenuated total reflection) and infrared microscopy on their application to measuring the same tissue for cancer diagnosis. The measurements showed that the coincidence of pathological diagnosis with ATR is higher than that of microscopy FTIR method. The result indicated that the measured area of tumor tissue is critical. The area for microscopy measurement is approximately 250 microm2, it reflects the heterogeneity of the tissue. However, the ATR measurement provides more information of malignant tissue for the measured area is 1.9 x 10(5) times than microscopy. Therefore, ATR exhibit the high accuracy in cancer detection for clinical application.

Fiber Optic Technology↗
PubMed1977

Scanning electron, light, and immunofluorescent microscopy of intestine of gnotobiotic calf infected with reovirus-like agent.

Intestinal lesions caused by a calf diarrheal reovirus-like agent were studied by scanning electron microscopy, light microscopy, and immunofluorescent microscopy in 2 gnotobiotic calves inoculated per orum. The calves were euthanatized 6 hours after the onset of diarrhea. Villi in the cranial and middle parts of the small intestine appeared shortened. In these areas, the epithelium was composed of low columnar and cuboidal cells. Villi in the caudal part of the small intestine were short. By scanning electron microscopy, the villous epithelium in the caudal part of the small intestine of both calves and the ridge epithelium in the ansa spiralis coli of 1 calf were composed of various sized cells which gave the surfaces an irregular appearance. The amount of mucopolysaccharide in the colonic epithelial cells of both calves was greatly reduced. The combined use of scanning electron and light microscopic techniques enabled obtaining a more accurate description of the intestinal lesions than either technique alone.

Animals↗
PubMed1990

Changes in the esophageal epithelium in rabbits treated by Cis-dichlorodiammineplatinum as studied by electron microscopy.

The esophageal mucosa of the rabbit was investigated after a single dose of 5 mg Cis-Dichlorodiammineplatinum (Cis-DDP). Specimens were taken for scanning electron microscopy, transmission electron microscopy, and light microscopy. Examination was performed daily for 20 consecutive days. A cytotoxic effect was observed already the first day after injection with an intracellular oedema. Thereafter the height of the esophageal epithelium and the basal cell layer steadily decreased to a minimum day 11. This parallels the damaged microridges and an increased cell loss as revealed by scanning electron microscopy. At the end of the observation period the esophageal mucosa had completely restituted.

Animals↗
PubMed1987

Scanning electron microscopy of cellular and acellular glomeruli of male dogs affected with Samoyed hereditary glomerulopathy and a carrier female.

Glomeruli isolated from three male dogs affected with Samoyed hereditary glomerulopathy were compared by scanning electron microscopy with glomeruli of one carrier female and six unaffected dogs. Scanning electron microscopy was performed before and after removal of podocytes and endothelial cells with enzyme and detergent, producing cellular and acellular glomeruli respectively. Cellular glomeruli of unaffected dogs showed podocytes with normally arranged foot processes, while in acellular glomeruli, the subepithelial surface of glomerular capillary basement membranes appeared smooth to finely granular. In contrast, cellular glomeruli of affected males showed microvilli, globular cytoplasmic projections from podocytes, and effacement of foot processes; acellular glomeruli demonstrated ridges and plaque-like irregularities on the subepithelial surface of glomerular capillary basement membranes. Changes in the glomeruli of the carrier female were intermediate between those of unaffected and affected male dogs. The appearance of the subepithelial surface of glomerular capillary basement membranes of acellular glomeruli seen by scanning electron microscopy correlated with the extent of multilaminar splitting of glomerular capillary basement membranes seen by transmission electron microscopy.

Animals↗
PubMed1987

Correlative light and electron microscopy of platelet adhesion and fibrinogen receptor expression using colloidal-gold labeling.

Differences in the shape change responses of platelets to various polymers may determine the thrombotic potential of these materials. Substrate-dependent variation in the expression and motility of the platelet fibrinogen receptor may underlie these differences due to this ligand's essential role in platelet aggregation. In this study we examine platelet activation on polyetherurethaneureas (PEUUs) presently being evaluated for vascular prosthetic applications. These polymers are prepared as 50-100nm thin films suitable as substrates for consecutive light microscopy, high voltage electron microscopy (HVEM), and SEM. 18nm colloidal gold coupled to fibrinogen permits visualization of that receptor's motility in living cells by video-enhanced light microscopy. Subsequent HVEM and SEM of identified cells provides correlative ultrastructure and surface morphology. The use of these novel support films coupled with the multiple modes of microscopy and colloidal gold labeled ligands permits in depth study of the molecular biology of cell adhesion to materials with varied, and known, surface properties. The motility of the platelet fibrinogen receptor was related to the extent of cytoskeletal reorganization, which, in turn, was influenced by polymer surface energetics. Platelets adherent to more hydrophobic PEUUs had greater receptor mobility and receptor redistribution than platelets adherent to more hydrophilic PEUUs. The most extensive receptor motility and redistribution was observed on Formvar, a non-PEUU with low surface-water energy, suggesting that additional surface properties are of importance in determining platelet spreading and fibrinogen receptor motility.

Biocompatible Materials↗
PubMed1979

Embedding in large plastic blocks. Diagnostic light and potential electron microscopy on the same block.

Specimens destined for light and electron microscopy were fixed in a modified buffered formalin, postosmicated, dehydrated, and embedded in a mixture of epoxy resins (Epon-araldite) in large plastic molds. These blocks were sectioned at 0.5 to 1 micron on a JB-4 microtome and stained with a combined nuclear and cytoplasmic stain (Paragon). The sections were examined by light microscopy for diagnostic evaluation. If ultrastructural examination was also desired, the selected area was isolated using the "mesa" technique. The trimmed block was then sectioned on an ultramicrotome, picked up on grids, stained, and examined in the electron microscope. We think these techniques offer the diagnostic pathologist the potential of viewing 1-micron sections at a light microscopy level with the option of subsequent electron microscopy of the same area of the same block.

Histological Techniques↗
PubMed1986

Analyses of phosphorylase kinase by transmission and scanning transmission electron microscopy.

Under conventional electron microscopy negatively stained phosphorylase kinase exhibits a bilobal structure resembling two bridged opposing parentheses. In this predominant particle orientation, usually only one bridge is observed; however, in many particles two bridges can be seen. Scanning transmission electron microscopy of unstained phosphorylase kinase shows very similar structures, with a particle mass equivalent to that of the hexadecameric holoenzyme. Partial digestion of the enzyme with chymotrypsin, which preferentially hydrolyzes the alpha-subunits, causes no significant changes in the structure; however, when both the alpha and beta subunits are degraded by trypsin, single lobed particles appear, i.e. the connecting bridges are missing. Mass analysis of scanning transmission electron microscopy images of trypsinized enzyme indicates that the protease does, in fact, split the particle into halves. Transmission electron microscopy of an alpha gamma delta complex isolated after incubation of the holoenzyme with LiBr shows only small particles approximately one-fourth the size of the holoenzyme. Thus, integrity of the beta subunit may be necessary in order for the two lobes of phosphorylase kinase to be bridged. These data also indicate that the subunits are arranged as a bridged dimer of octamers 2 (alpha 2 beta 2 gamma 2 delta 2).

Animals↗
PubMed1983

Carbohydrate cytochemistry by transmission and scanning electron microscopy.

Macromolecules containing abundant carbohydrate moieties occur widely distributed on cell surfaces and in cell organelles. Extension to the ultrastructural level of light microscopic cytochemical methods for demonstrating the complex carbohydrates has provided extensive information about the precise location of a great diversity of glycoconjugates in various cell types affording, accordingly, an improved insight into the biologic significance of the chemical nature of some components. The available methods of ultrastructural carbohydrate cytochemistry are reviewed in this tutorial, and the applicability of these methods to demonstration of glycoconjugates by scanning electron microscopy is considered. Backscattered electron imaging was carried out on specimens of trachea stained by methods for demonstrating glycoconjugates by transmission electron microscopy. This approach provided a means of localizing sites of abundant complex carbohydrate by scanning electron microscopy. Further extension of carbohydrate cytochemical methods to demonstration of glycoconjugates by scanning electron microscopy appears feasible and offers a promise of providing valuable information for carbohydrate cytochemistry.

Animals↗
PubMed1982

[Modern electron microscopy methods in the study of ultrastructure and function of the terminal vascular system].

The present survey deals with the possibilities of electron microscopic methods, with the gain of information by its use and with its prospective significance in solving actual problems in the subject of microcirculation. The paper consists of the subsequent parts: 1. Fixation methods; 2. Topical methods for ultrastructural investigation of defined areas of the terminal vascular bed and the transmural ways of transport of molecules; 3. Freeze etching technics; 4. Electron microscopic autoradiography; 5. Transmission electron microscopy including methods for three-dimensional reconstruction of vascular segments; 6. Scanning electron microscopy; 7. X-ray microanalysis; 8. Demonstration of cell receptors with the aid of lectines; 9. Methods for quantitative analysis. In each chapter the possibilities and disadvantages of the methods and the most frequently occurring artefacts were dealt. The survey concludes with a consideration of the prospective development of electron microscopic technics and its use in the investigation of the terminal vascular bed. In the opinion of the authors the methodical development will take the subsequent direction: 1. Further completion of the preparative methods and its development to routine technics combined with the more general use of necessary tools; 2. Further completion and spreading of quantitative investigation methods; 3. More effective use of the synthetic analysis solving a problem, that means, the simultaneous application of different methods (for instance: transmission electron microscopy, scanning electron microscopy, experimental methods and so on).

Animals↗