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At least 1,405 records · Page 78Linked to original sources
PubMed2004

Viability of endolithic micro-organisms in rocks from the McMurdo Dry Valleys of Antarctica established by confocal and fluorescence microscopy.

The rocks of the McMurdo Dry Valleys desert in Antarctica harbour endolithic communities of micro-organisms such as lichens, fungi, cyanobacteria and bacteria. Establishing the physiological status and viability of these microbial colonies in their natural microhabitat has far-reaching implications for understanding the microbial ecology of the harsh environment of this polar desert. Here we describe the use of confocal microscopy and a specific fluorescent probe (FUN-1) to evaluate the metabolic activity of fungal cells. Application of confocal microscopy also served to identify living and dead bacteria or cyanobacteria using the fluorescent assay reagents Live/Dead SYTO 9 and propidium iodide or SYTOX Green, respectively. In addition, through the use of epifluorescence microscopy, live/dead bacteria and cyanobacteria could be detected by estimating fluorescence from their cell components provoked by simultaneously staining with nucleic acids stains such as DAPI and SYTOX Green.

Antarctic Regions↗
PubMed1984

Colonial morphology of treponemes observed by electron microscopy.

The colonial morphology of three strains of cultivable, nonpathogenic treponemes including a human oral treponeme was examined by light and electron microscopy. Treponema phagedenis strains Kazan and Reiter produced large white colonies on the surface of solid media composed of sterility test broth, 0.9 to 3.1% agar, rifampin, and 12.5% rabbit or horse serum. A human oral treponeme, strain G7201 , grew as diffused white zones on 0.9 to 3.1% agar plates. Under the cultural conditions employed agar concentrations slightly affected the time of appearance of colonies of the three strains of treponemes. When the colonies of these three strains were viewed by scanning electron microscopy, differences in their colonial morphology were observed. The 11-day-old colonies of human oral strain G7201 were very small, 5 to 15 micron in diameter, and had a slight irregular border. Kazan treponemes developed circular, entire and low convex colonies. Scanning and transmission electron microscopy revealed that the colonies of Reiter treponemes contained spherical forms almost up to 5 micron in diameter, each consisting of an outer membrane and a treponemal main body. They were very similar to the spherical bodies produced by strain G7201 in sucrose-containing broth.

Agar↗
PubMed1978

Differential leucocyte counts: a comparison of results using light and electron microscopy.

A method is described for handling leucocytes from an inflammatory peritoneal exudate prior to electron microscopy, which allows differential counts from ultra-thin sections to be made. The results of counts from ultrathin sections, viewed on the electron microscope, are compared with samples from the same cell populations prepared on a cytocentrifuge and counted by light microscopy. The results from several cell populations of widely different compositions show clearly that with suitable care taken over preparation and orientation, ultrathin sections can yield comparable differential counts to those obtained by standard light microscope procedures. Possible sources of error are discussed and the advantages of ultrastructural counting assessed. The method has a wide application wherever accurate differential counts are required from cell suspensions processed for electron microscopy.

Ascitic Fluid↗
PubMedDate not supplied

Tissue response in sporotrichosis: light and electron microscopy studies.

On the basis of electron microscopy examination of human tissue affected by sporotrichosis, we have proposed a classification, at the light microscopy level, of fungal cells in tissue. Light and electron microscopy observations clarified the following five points: (1) in sporotrichosis lesions in man, the fungal cells are frequently phagocytized by polymorphonuclear leucocytes (PMNs); (2) phagocytosis of phagocytes by other phagocytes was common, and giant cells phagocytized both fungal cells and PMNs; (3) fungal cells in the tissues of lesions were thought to be highly viable, and their budding was frequently observed within macrophages and giant cells; (4) it was highly probable that PMNs participated in asteroid body formation; (5) few free fungal cells were present in the tissues. We believe that the classification of fungal cells in sporotrichosis tissue into the five categories below is useful for correctly understanding the pathological condition in a sporotrichosis lesion: (1) fungal cells in PMNs, (2) fungal cells in PMNs within macrophages, (3) fungal cells in macrophages, (4) fungal cells in giant cells, and (5) free fungal cells.

Giant Cells↗
PubMed2006

Cell signaling and trafficking of human melanocortin receptors in real time using two-photon fluorescence and confocal laser microscopy: differentiation of agonists and antagonists.

Melanocortin hormones and neurotransmitters regulate a vast array of physiologic processes by interacting with five G-protein-coupled melanocortin receptor types. In the present study, we have systematically studied the regulation of individual human melanocortin receptor wild subtypes using a synthetic rhodamine-labeled human melanotropin agonist and antagonist, arrestins fused to green fluorescent protein in conjunction with two-photon fluorescence laser scanning microscopy and confocal microscopy. Stimulation of the melanocortin receptors by its cognate agonist triggered rapid arrestin recruitment and receptor internalization for all four human melanocortin receptors examined. Antagonists-bound melanocortin receptors, on the other hand, did not recruit beta-arrestins, and remained in the cell membrane even after long-term (30 min) treatment. Agonist-mediated internalization of all melanocortin receptor subtypes was sensitive to inhibitors of clathrin-dependent endocytosis, but not to caveolae inhibitors. In summary, agonist-mediated internalization of all subtypes of melanocortin receptors are dependent upon beta-arrestin-mediated clathrin-coated pits, whereas, beta-arrestin-2 conjugated green fluorescence protein (beta-arrestin-2-GFP) recruitment is not dependent on protein kinase A activation. Real time two-photon fluorescence laser scanning microscopy is a most powerful tool to study the dynamic processes in living cells and tissues, without inflicting significant and often lethal damage to the specimen.

Arrestins↗
PubMed2001

Blood cell adhesion on sensor materials studied by light, scanning electron, and atomic-force microscopy.

Unwanted interactions of biomedical sensors with surrounding tissues, body fluids, and cells are one of the most crucial problems affecting their long-term stability. In vivo processes were simulated in a computer-controlled bioreactor connected to a flow chamber system. Optical sensor materials were inserted into a parallel-plate chamber and monitored by light microscopy in order to get information about the number of adhered cells. Tests with thrombocyte-enriched plasma show that novel phosphorylcholine (PC)-polymer-coated sensors appear to be more bioinert, and thus demonstrate better haemocopatibility in comparison with untreated glass sensors. The influence of different materials on the morphology of adhered cells was investigated by off-line methods such as scanning electron microscopy (SEM) and atomic-force microscopy (AFM). SEM showed a reduction in the number of adhered thrombocytes and the lack of any fibrin network on the PC-polymer-modified glass surface, in contrast to the pure glass surface. AFM gives topographical information, and the calculated contact areas and cell volumes indicate smaller interactions between cells and sensor material in the case of PC-polymer-coated sensors.

Bioreactors↗
PubMed2001

Second-harmonic imaging microscopy of living cells.

Second harmonic generation (SHG) has been developed in our laboratories as a high-resolution nonlinear optical imaging microscopy for cellular membranes and intact tissues. SHG shares many of the advantageous features for microscopy of another more established nonlinear optical technique: two-photon excited fluorescence (TPEF). Both are capable of optical sectioning to produce three-dimensional images of thick specimens and both result in less photodamage to living tissue than confocal microscopy. SHG is complementary to TPEF in that it uses a different contrast mechanism and is most easily detected in the transmitted light optical path. It can be used to image membrane probes with high membrane specificity and displays extraordinary sensitivity in reporting membrane potential; it also has the ability to image highly ordered structural proteins without any exogenous labels.

Animals↗
PubMed2003

Analysis of histology specimens using lifetime multiphoton microscopy.

Observations of cells or tissues with fluorescence microscopy can provide unique insights into cellular physiology and structure. Such information may reveal the pathological state of a tissue to the physician or information on cytoskeletal dynamics to the research scientist. However, problems of overlapping spectra, low signal, and light scatter impose serious limitations on what can be achieved in practice with fluorescence microscopy. These problems can be addressed in part by the development of new imaging modalities that make maximum use of the information present in the fluorescence signal. We describe the application of a new technology to the study of standard histological pathology specimens: a multiphoton excitation fluorescence microscope that incorporates a novel, photon-counting detector that measures the excited-state lifetimes of fluorescent probes. In initial investigations, we have applied this system to the observation of C. elegans embryos and primate histology specimens, with the objective of identifying potentially diagnostic signatures. Our findings demonstrate that lifetime multiphoton microscopy has considerable potential as a diagnostic tool for pathological investigations.

Animals↗
PubMedDate not supplied

Excitation beyond the monochromatic laser limit: simultaneous 3-D confocal and multiphoton microscopy with a tapered fiber as white-light laser source.

Confocal and multiphoton microscopy are essential tools in modern life sciences. They allow fast and highly resolved imaging of a steadily growing number of fluorescent markers, ranging from fluorescent proteins to quantum dots and other fluorophores, used for the localization of molecules and the quantitative detection of molecular properties within living cells and organisms. Up to now, only one physical limitation seemed to be unavoidable. Both confocal and multiphoton microscopy rely on lasers as excitation sources, and their monochromatic radiation allows only a limited number of simultaneously usable dyes, which depends on the specific number of laser lines available in the used microscope. We have overcome this limitation by successfully replacing all excitation lasers in a standard confocal microscope with pulsed white light ranging from 430 to 1300 nm generated in a tapered silica fiber. With this easily reproducible method, simultaneous confocal and multiphoton microscopy was demonstrated. By developing a coherent and intense laser source with spectral width comparable to a mercury lamp, we provide the flexibility to excite any desired fluorophore combination.

Animals↗
PubMed1998

Electron and fluorescence microscopy of extracellular glucan and aryl-alcohol oxidase during wheat-straw degradation by Pleurotus eryngii.

The ligninolytic fungus Pleurotus eryngii grown in liquid medium secreted extracellular polysaccharide (87% glucose) and the H2O2-producing enzyme aryl-alcohol oxidase (AAO). The production of both was stimulated by wheat-straw. Polyclonal antibodies against purified AAO were obtained, and a complex of glucanase and colloidal gold was prepared. With these tools, the localization of AAO and extracellular glucan in mycelium from liquid medium and straw degraded under solid-state fermentation conditions was investigated by transmission electron microscopy (TEM) and fluorescence microscopy. These studies revealed that P. eryngii produces a hyphal sheath consisting of a thin glucan layer. This sheath appeared to be involved in both mycelial adhesion to the straw cell wall during degradation and AAO immobilization on hyphal surfaces, with the latter evidenced by double labelling. AAO distribution during differential degradation of straw tissues was observed by immunofluorescence microscopy. Finally, TEM immunogold studies confirmed that AAO penetrates the plant cell wall during P. eryngii degradation of wheat straw.

Alcohol Oxidoreductases↗
PubMed2003

Scanning transmission X-ray, laser scanning, and transmission electron microscopy mapping of the exopolymeric matrix of microbial biofilms.

Confocal laser scanning microscopy (CLSM), transmission electron microscopy (TEM), and soft X-ray scanning transmission X-ray microscopy (STXM) were used to map the distribution of macromolecular subcomponents (e.g., polysaccharides, proteins, lipids, and nucleic acids) of biofilm cells and matrix. The biofilms were developed from river water supplemented with methanol, and although they comprised a complex microbial community, the biofilms were dominated by heterotrophic bacteria. TEM provided the highest-resolution structural imaging, CLSM provided detailed compositional information when used in conjunction with molecular probes, and STXM provided compositional mapping of macromolecule distributions without the addition of probes. By examining exactly the same region of a sample with combinations of these techniques (STXM with CLSM and STXM with TEM), we demonstrate that this combination of multimicroscopy analysis can be used to create a detailed correlative map of biofilm structure and composition. We are using these correlative techniques to improve our understanding of the biochemical basis for biofilm organization and to assist studies intended to investigate and optimize biofilms for environmental remediation applications.

Bacteria↗
PubMed1969

Scanning electron and phase-contrast microscopy of bacterial spores.

The three-dimensional immages of free and intrasporangial spores produced by scanning electron microscopy show surface structures not visible by phase-contrast microscopy. Although fine surface detail is not elucidated by scanning electron microscopy, this technique does afford a definitive picture of the general shape of spores. Spores of Bacillus popilliae, B. lentimorbus, B. thuringiensis, B. alvei, B. cereus, and Sarcina ureae have varying patterns of surface ridge formation, whereas spores of B. larvae, B. subtilis, and B. licheniformis have relatively smooth surfaces.

Bacillus↗
PubMed2004

Atomic force microscopy of cell growth and division in Staphylococcus aureus.

The growth and division of Staphylococcus aureus was monitored by atomic force microscopy (AFM) and thin-section transmission electron microscopy (TEM). A good correlation of the structural events of division was found using the two microscopies, and AFM was able to provide new additional information. AFM was performed under water, ensuring that all structures were in the hydrated condition. Sequential images on the same structure revealed progressive changes to surfaces, suggesting the cells were growing while images were being taken. Using AFM small depressions were seen around the septal annulus at the onset of division that could be attributed to so-called murosomes (Giesbrecht et al., Arch. Microbiol. 141:315-324, 1985). The new cell wall formed from the cross wall (i.e., completed septum) after cell separation and possessed concentric surface rings and a central depression; these structures could be correlated to a midline of reactive material in the developing septum that was seen by TEM. The older wall, that which was not derived from a newly formed cross wall, was partitioned into two different surface zones, smooth and gel-like zones, with different adhesive properties that could be attributed to cell wall turnover. The new and old wall topographies are equated to possible peptidoglycan arrangements, but no conclusion can be made regarding the planar or scaffolding models.

Cell Division↗
PubMed1965

Phase-contrast and electron microscopy of murine strains of Mycoplasma.

Nelson, John B. (The Rockefeller University, New York, N.Y.), and Michael J. Lyons. Phase-contrast and electron microscopy of murine strains of Mycoplasma. J. Bacteriol. 90:1750-1763. 1965.-Two strains of Mycoplasma pulmonis (associated with infectious catarrh) on examination in fluid culture (20% horse serum-bouillon) by phase microscopy were highly pleomorphic, with many bacilliform elements and fewer coccoid ones. Motility, characterized by gliding of rods and spinning of spherical forms, was observed through the 9th subculture of one strain and the 15th of the second. Motile elements were not seen in later transfers and pleomorphism was reduced. One strain of M. neurolyticum (associated with conjunctivitis and encephalitis) was much less pleomorphic and showed neither bacilliform elements nor motility at any time. When examined by negative-contrast electron microscopy, organisms of this strain were found to have an average diameter of 0.7 mu and to possess a concentrated peripheral layer of cytoplasm and a central mass which may represent the cells' nuclear equivalent. The latter feature was not prominent in spherical forms of M. pulmonis. These cells, when observed after 48 hr of culture, showed evidence of the generation of new progeny cells in their central area. The filamentous or bacilliform cells of M. pulmonis were frequently serpentine in appearance, 2.0 to 3.0 mu in length and 80 to 250 mmu in width. They appeared to generate new cells from terminal buds from which outpouchings initially developed. Older cells, in the stationary phase, showed evidence of undergoing multipolar germination. Microtubules, about 60 mmu wide, were found in association with most filamentous cells from 48-hr cultures; fragments of membrane, studded with closely packed ribosomelike particles, were also found. There was no evidence of flagella or any specialized structure that could account for the observed motility of the organisms.

Animals↗
PubMed2005

Contribution to comprehension of image formation in confocal microscopy of cornea with Rostock cornea module.

AIM: To investigate the influence of refractive index of aqueous humour on imaging of corneal endothelium in confocal microscopy. To clarify the phenomenon of dark endothelial and bright epithelial cell membranes in confocal images of corneas. METHODS: Use of a novel digital confocal laser scanning microscope, a combination of the Heidelberg retina tomograph (HRT II) and the Rostock cornea module. Exchange of aqueous humour solution from domestic pigs against glycerol/water solutions (refractive indices eta = 1.337-1.47). Transelectron microscopy of endothelial and epithelial cell morphology. RESULTS: Under the terms of variable refractive indices no differences were observed for general imaging of endothelium. Bright cells were bordered by dark cell membranes in all experiments. Electron microscopy of endothelium and epithelium revealed differences in intracellular and cell membrane structure of both cell types. CONCLUSION: Source of specific confocal optical behaviour of endothelium does not come from interface conditions to aqueous humour, but may result from intracellular variations and ultrastructure of cell membranes.

Animals↗
PubMed1977

The sporulation process in Thermomonospora fusca as revealed by scanning and transmission electron microscopy.

The sporulation process in the thermophilic actinomycete Thermomonospora fusca was observed by scanning and transmission electron microscopy. As shown by scanning electron microscopy, spores were produced primarily on aerial hyphae and first appeared as bud-like enlargements at the tips of short multibranched sporophores. Young spores were oval to spherical in shape with a smooth surface. As they matured spores enlarged and developed a rough and globular covering, which was quite fragile and easily detached from the spore. This outer layer, as observed by transmission electron microscopy, was thought equivalent to the sheath of other Thermomonospora species. In cross section, mature spores exhibited a thick spore coat underneath the outer globular layer. This spore coat was usually observed as a single layer, but some spores produced a bilayered coat. No multilayered spore coat or spore cortex was observed in the heat-sensitive spores of T. fusca. They were, therefore, shown to be aleuriospores (microcondia), and not endospores.

Cell Wall↗
PubMed1989

Use of electron microscopy to characterize the surfaces of flocculent and nonflocculent yeast cells.

The surfaces of flocculent and nonflocculent yeast cells have been examined by electron microscopy. Nonextractive preparative procedures for scanning electron microscopy allow comparison in which sharp or softened images of surface details (scars, etc.) are the criteria for relative abundance of flocculum material. Asexually flocculent budding-yeast cells cannot be distinguished from nonflocculent budding-yeast cells in scanning electron micrographs because the scar details of both are well resolved, being hard and sharp. On the other hand, flocculent fission-yeast cells are readily distinguished from nonflocculent cells because fission scars are mostly soft or obscured on flocculent cells, but sharp on nonflocculent cells. Sexually and asexually flocculent fission-yeast cells cannot be distinguished from one another as both are heavily clad in "mucilaginous" or "hairy" coverings. Examination of lightly extracted and heavily extracted flocculent fission-yeast cells by transmission electron microscopy provides micrographs consistent with the scanning electron micrographs.

Flocculation↗