Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Fluorescence Microscopy”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 883 records · Page 49Linked to original sources
PubMed1999

Modulation of mitochondrial respiration by nitric oxide: investigation by single cell fluorescence microscopy.

With the electro-driven import of rhodamine 123, we used single cell fluorescence microscopy to single out the contribution of nitric oxide (NO) in controlling mitochondrial membrane potential expressed by (stationary growing) rhabdomyosarcoma and neuroblastoma cells in culture. The experimental design and the computer-aided image analysis detected and quantitated variations of fluorescence signals specific to mitochondria. We observed that 1) the two cell lines display changes of fluorescence dependent on mitochondrial energization states; 2) mitochondrial fluorescence decreases after exposure of the cells to a NO releaser; 4) the different fluorescence intensity measured under stationary growing conditions, or after activation and inhibition of constitutive NO synthase, is consistent with a steady-state production of NO. Direct comparison of single cell fluorescence with bulk cytofluorimetry proved that the results obtained by the latter method may be misleading because of the intrinsic-to-measure lack of information about distribution of fluorescence within different cell compartments. The kinetic parameters describing the reactions between cytochrome oxidase, NO, and O2 may account for the puzzling (20-fold) increase of the KM for O2 reported for cells and tissues as compared to purified cytochrome c oxidase, allowing an estimate of in vivo NO flux.

Fluorescence↗
PubMed1980

Immunoperoxidase methods: increased efficiency using fluorescence microscopy for 3,3-diaminobenzidine (DAB) stained semithin sections.

When semithin sections stained by immunoperoxidase-DAB methods are exposed to ultraviolet light in a fluorescence microscope, immunoreactive cells develop a strong yellowish fluorescence within 2-4 min. This property offers the possibility of visualizing even reaction products which can barely be identified by other microscopical techniques. Thus the efficiency of immunoperoxidase methods is greatly enhanced. Moreover the histochemical proof of endogenous peroxide active enzymes visualized with DAB as substrate may also be facilitated using fluorescence microscopy.

3,3'-Diaminobenzidine↗
PubMed1988

The rapid desensitization of receptors for platelet derived growth factor, bradykinin and ATP: studies on individual cells using quantitative digital video fluorescence microscopy.

The rise in free cytosolic Ca2+ of individual response to growth factors was studied in serum starved cultures of 3T3 fibroblasts. Quantitative digital video fluorescence microscopy revealed that with platelet derived growth factor (PDGF) there was a lag period between stimulation and Ca2+ response, with considerable cell-to-cell variation, whereas ATP, bradykinin and fetal calf serum induced an immediate, synchronous response. A coverslip with attached cells was mounted on a small flow chamber, allowing complete change of medium in 2 sec. Using this technique, homologous desensitization to a second addition of agonist 2 min after removal of the first addition was found for all agonists. Unusual heterologous desensitization was observed in that PDGF desensitized the cells to the other agonists, yet the reverse did not occur.

Adenosine Triphosphate↗
PubMedDate not supplied

[The viability assessment of ethanol-producing yeast by computer-aided fluorescence microscopy].

Vital staining of the ethanol-producing yeast Saccharomyces cerevisiae with ethidium bromide and DAPI allows intact and damaged cells to be differentiated by fluorescence microscopy. A computer image analysis procedure has been developed for the automatic determination of the relative number of damaged cells using ImageJ software (National Institute of Health, United States; http://rsb.info.nih.gov./ij/). A good correlation has been found between the viability rates determined by the plate count method and the relative numbers of intact cells assessed by the developed procedure in the dry-yeast preparations rehydrated under various conditions.

Colony Count, Microbial↗
PubMed1994

In vivo fluorescence microscopy of microcirculation in the renal cortex of mice. Part V. Effects of mannitol and iohexol infusions in normal, obese/hyperglycemic and diabetic mice.

In vivo fluorescence microscopy was used in experimental studies of renal cortical microcirculation in mice. The effects of i.v. infusions of mannitol and iohexol were studied in normal and obese/hyperglycemic mice and in mice with streptozotocin-induced diabetes mellitus. All infusions produced marked effects on the distribution and velocity of cortical blood flow with no differences between iohexol and mannitol. The renal cortical blood flow was inhomogeneous regarding different capillaries and there was an initial rise in the number of capillaries with increased blood flow after infusions of both mannitol and iohexol. The infusions of contrast medium were initially followed by a significantly larger number of capillaries with decreased blood flow in the obese/hyperglycemic mice (p < 0.05) than in the diabetic and nondiabetic mice. The variations in obese/hyperglycemic groups were, however, large. The effects were the same in normal and diabetic animals.

Animals↗
PubMed1997

Far-field fluorescence microscopy with three-dimensional resolution in the 100-nm range.

We report three-dimensional (3D) microscopy with nearly isotropic resolution in the lambda/5-lambda/10 range. Our approach combines 4PI-confocal two-photon fluorescence microscopy with image restoration. The 3D resolution is demonstrated with densely clustered beads as well as with F-actin fibers in mouse fibroblast cells. A comparison with unrestored two-photon confocal images reveals a total reduction of the uncertainty volume up to a factor of 15.

Animals↗
PubMed1992

Dynamical conformational change of DNA induced by synthetic polymers: direct observation by fluorescence microscopy.

Brownian motion and Higher-order structure of DNA molecules complexed with synthetic polymers: poly-L-arginine (pArg) and polyacrylamide (pAAm), were observed using fluorescence microscopy. Quantitative analyses of the conformation and motion of the complexes demonstrate that these two polymers influence the structure of DNA molecules in an opposite way. The electrostatic binding of pArg induces contraction of DNA, whereas the intercalative pseudo-grafting of pAAm extends DNA.

Acrylic Resins↗
PubMed2003

Three-dimensional optical-transfer-function analysis of fiber-optical two-photon fluorescence microscopy.

The three-dimensional optical transfer function is derived for analyzing the imaging performance in fiber-optical two-photon fluorescence microscopy. Two types of fiber-optical geometry are considered: The first involves a single-mode fiber for delivering a laser beam for illumination, and the second is based on the use of a single-mode fiber coupler for both illumination delivery and signal collection. It is found that in the former case the transverse and axial cutoff spatial frequencies of the three-dimensional optical transfer function are the same as those in conventional two-photon fluorescence microscopy without the use of a pinhole.However, the transverse and axial cutoff spatial frequencies in the latter case are 1.7 times as large as those in the former case. Accordingly, this feature leads to an enhanced optical sectioning effect when a fiber coupler is used, which is consistent with our recent experimental observation.

Journal Article↗
PubMed2003

Computational analysis of F-actin turnover in cortical actin meshworks using fluorescent speckle microscopy.

Fluorescent speckle microscopy (FSM) is a new imaging technique with the potential for simultaneous visualization of translocation and dynamic turnover of polymer structures. However, the use of FSM has been limited by the lack of specialized software for analysis of the positional and photometric fluctuations of hundreds of thousand speckles in an FSM time-lapse series, and for translating this data into biologically relevant information. In this paper we present a first version of a software for automated analysis of FSM movies. We focus on mapping the assembly and disassembly kinetics of a polymer meshwork. As a model system we have employed cortical F-actin meshworks in live newt lung epithelial cells. We lay out the algorithm in detail and present results of our analysis. The high spatial and temporal resolution of our maps reveals a kinetic cycling of F-actin, where phases of polymerization alternate with depolymerization in a spatially coordinated fashion. The cycle rates change when treating cells with a low dose of the drug latrunculin A. This shows the potential of this technique for future quantitative screening of drugs affecting the actin cytoskeleton. Various control experiments demonstrate that the algorithm is robust with respect to intensity variations due to noise and photobleaching and that effects of focus plane drifts can be eliminated by manual refocusing during image acquisition.

Actins↗
PubMed2002

Spectral imaging fluorescence microscopy.

The spectral resolution of fluorescence microscope images in living cells is achieved by using a confocal laser scanning microscope equipped with grating optics. This capability of temporal and spectral resolution is especially useful for detecting spectral changes of a fluorescent dye; for example, those associated with fluorescence resonance energy transfer (FRET). Using the spectral imaging fluorescence microscope system, it is also possible to resolve emitted signals from fluorescent dyes that have spectra largely overlapping with each other, such as fluorescein isothiocyanate (FITC) and green fluorescent protein (GFP).

Fluorescein-5-isothiocyanate↗
PubMed1986

A simple method to select specific tumor areas in paraffin blocks for cytometry using incident fluorescence microscopy.

A simple method is described for the selection of tumor areas in paraffin blocks for cytometry. The surface of a paraffin block is stained with the fluorescent dye DAPI. By means of incident fluorescence microscopy on the cut surface of a total block, the tissue can be visualized. Location of the tumor area with the aid of conventional histopathological criteria is feasible using the adjacent section after hematoxylin-eosin staining as a diagnostic guideline. Once the position of the tumor area is determined, a small hollow bore, which is screwed in the objective holder, is pressed in the tissue. The depth is controlled by the microscrew of the microscope. After retraction of the bore, the block is removed, and a thick section is cut. The selected area can be processed for cytometry separately from the remaining tissue. The technique can be used either to enrich the sample to be analyzed with tumor cells or to analyze histopathologically different tumor compartments. Both flow and image cytometry can make use of this selection technique.

Breast↗
PubMed1991

Confocal fluorescence microscopy and three-dimensional reconstruction.

Several recent technological advances have considerably improved the field of confocal fluorescence microscopy. Improvements in confocal microscope design, new fluorescent probes and indicators, more sensitive imaging devices, and computer advances which allow for data manipulation and storage provide a convenient method to acquire complex three-dimensional (3-D) architectural details which previously were difficult or impossible to obtain from biological specimens. Applications of the laser scanning and tandem scanning confocal microscopes offer the potential for gaining powerful insights into the complex relationship of cellular structure and function. Confocal microscopy generates optical sections free from out-of-focus blur. With the development of new visualization tools to render and display complex 3-D data, a set of optical sections taken at different focal planes can be three-dimensionally reconstructed to create an animated sequence which can reveal latent features of the specimen. The combination of confocal microscopy and 3-D reconstruction provides a powerful new imaging tool to advance knowledge about structural and functional cellular properties as they occur dynamically in three dimensions.

Animals↗
PubMed1989

Calcium-triggered movement of regulated actin in vitro. A fluorescence microscopy study.

We previously reported setting up an in vitro system for the observation of actin filament sliding along myosin filaments. The system involved a minute amount of fluorescently labelled F-actin, and its movement was monitored by fluorescence microscopy. Here, we report observations of the Ca2+-dependent movement of F-actin complex with tropomyosin plus troponin (regulated actin) added to the movement system in place of pure F-actin. In a wide range of pCa (-log10[Ca2+]) between 3 and 5.5 at 30 degrees C, regulated actin filaments moved rapidly, and the average velocity depended little on the Ca2+ concentration (about 7.5 microns/s). However, when the Ca2+ concentration was decreased to pCa = 5.8 or lower, the filaments suddenly stopped moving. In striking contrast to these observations, unregulated actin moved rapidly within the whole pCa range examined, the average velocity (about 7.5 microns/s) being essentially Ca2+-independent. These observations indicate that (1) tropomyosin-troponin actually gave Ca2+-sensitivity to F-actin, and (2) the movement system was regulated by Ca2+ in an on-off fashion within a narrow range of Ca2+ concentration. In a pCa range between 5.8 and 6.0, regulated actin filaments did not exhibit thermal motion; instead, they had fixed positions in the specimen, possibly because they remained associated with myosin filaments in the background, without sliding past each other. Although regulated actin moved fast in the presence of 1 mM-CaCl2 (pCa = 3) at 30 degrees C, it became entirely non-motile as the temperature was decreased to 25 degrees C or lower. Such a sharp movement/temperature relation was never found for unregulated actin. We assayed regulated actin-activated myosin ATPase in the same conditions as used for microscopy, and found that the ATPase activity depended both on pCa and on the temperature considerably less than the movement of regulated actin. The results suggest that the sliding velocity in the in vitro system would not be proportional to the rate of actin-activated ATPase.

Actins↗
PubMed1997

Effects of gamma-aminobutyric acid, progesterone and ionophore A23187 on acrosome reaction of tree shrew sperm in vitro: examination of acrosome reaction with an improved fluorescence microscopy.

A number of acrosome reaction (AR) initiators have been found to be effective in inducing AR of human, laboratory and domestic animal sperm. Using an improved simple fluorescence microscopy, effects of gamma-aminobutyric acid (GABA), progesterone and ionophore A23187 on sperm AR of tree shrew, a useful animal model in biomedical research, have been investigated. Spontaneous AR in 4.92-7.53% of viable sperm was observed. Complete AR in 10.31-18.25% of viable tree shrew sperm was obviously induced by 5 microM and 10 microM calcium ionophore A23187, 1 mM GABA, and 5 microM progesterone, and there were no significant differences between their abilities to initiate complete AR. No significant differences of AR percentages between 1- and 2-h treatments with A23187, progesterone and/or GABA were observed. These results suggested that the responses of tree shrew sperm to these AR initiators are similar to that of human and other mammalian sperm.

Acrosome↗
PubMedDate not supplied

Localization of adhesion molecules on human spermatozoa by fluorescence microscopy.

The expression of adhesion molecules on human spermatozoa of healthy probands was analysed. The localization patterns of adhesion molecules (AM) on the spermatozoal surface were documented by fluorescence microscopy. Spermatozoa were incubated with antibodies against alpha 1 (CD49a), alpha 2 (CD49b), alpha 3 (CD49c), alpha 4 (CD49d), alpha 5 (CD49e), alpha 6 (CD49f) chains of beta 1 integrins, beta 1 (CD29), beta 2 (CD18), alpha V (CD51), beta 3 (CD61) and beta 4 integrin chains, the LFA-3 (Lymphocyte function antigen, CD58) from the immunoglobulin superfamily and the extra-cellular matrix proteins laminin, fibronectin and collagen IV. For collagen IV, alpha 1 and alpha 2 chains no expression could be noticed. Laminin was detected at the acrosomal membrane, fibronectin and beta 4 chain mainly at the equatorial membrane. The fibronectin receptors alpha 3, alpha 4 and alpha 5 chains of the beta 1 integrins were mainly located on acrosomal and equatorial membrane areas. Laminin receptor alpha 6 chain was located postacrosomal and less frequently acrosomal. beta 2 chain and vitronectin receptors alpha V and beta 3 chains had a mainly postacrosomal localization pattern. LFA-3 was found constantly on postacrosomal membrane areas. Double staining technique was used to prove the simultaneous occurrence of fibronectin and its integrin receptors alpha 3, alpha 4 and alpha 5 chains and of alpha V and beta 2 chains on spermatozoa. The localization patterns of integrins on double stained spermatozoa were similar to the patterns described for single stained spermatozoa. The localization of fibronectin appeared to be influenced by the presence of integrins: the typical equatorial fibronectin band disappeared in case of an equatorial localization of integrins.

Antigens, CD↗
PubMed1980

[Use of fluorescence microscopy in the cytochemical evaluation of wound healing after gingivectomy].

The authors investigated in 60 cases the healing process after gingivectomy. The investigation was performed by means of fluorescence microscopy, the presence of nucleic acids (RNA, DNA) and of polysaccharides in the cells being demonstrated. The cytochemical RNA reactions were the most sensitive indicator of wound healing after gingivectomy. Solcoseryl gel accelerated the epithelization of the gingival wound after the surgical intervention, which shortens the period of recuperation.

Adult↗
PubMed1980

Distribution of albumin in the normal monkey eye as revealed by Evans blue fluorescence microscopy.

Since intravenously injected Evans blue binds irreversibly to serum albumin, its distribution reflects albumin exchange between the intravascular and extravascular tissue compartments. In histologic specimens examined by fluorescence, microscopy, extravasated Evans blue--albumin complex was identified within the ciliary body and trabecular meshwork of normal monkey eyes. In eyes fixed by intra-arterial perfusion of fixative, no dye was identified in the choroid, retina, or optic nerve. With immersion fixation, however, some extravasation was seen in the choroid and adjacent optic nerve. In some specimens, the optic nerve was stained not only with material apparently leaking from the choroid but also from a breakdown of the blood-brain barrier in the major disc vasculature during the interval before fixative penetrates into the tissue. Perfusion fixation must be used to avoid this artifact, and freezing techniques would be even better.

Animals↗