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Results for “Microscopy, Confocal”

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At least 1,117 records · Page 62Linked to original sources
PubMed1992

Interphotoreceptor retinoid binding protein induced experimental autoimmune uveitis: an immunophenotypic analysis using alkaline phosphatase anti-alkaline phosphatase staining, dual immunofluorescence and confocal microscopy.

Using a Lewis rat model of IRBP induced EAU, we have examined the progress of leucocytes infiltrating the uveitic eye. APAAP and dual immunofluorescence were used to show that ED7 and 8 (CD11b/CD18) positive monocytes, W3/25 and OX8 (CD4 and CD8) lymphocytes were prominent in the initial inflammatory exudate around the retinal vessels and in the choroid. ED1 positive monocytes were also observed in the choroid. As disease progressed, these cells moved into the inner retina, vitreous and ROS. ED8 positive cells were the first to appear in the ROS. This was followed by the later appearance of ED2 tissue macrophages in the vitreous and ED3 inflammatory macrophages in the vitreous and ROS.

Alkaline Phosphatase↗
PubMedDate not supplied

Tandem-scanning (confocal) microscopy of the full-thickness cornea.

We have utilized a radically new type of optical scanning microscope to study the full-thickness morphology of the intact cornea in an excised human eye bank eye and in freshly sacrificed rabbit eyes in situ. This technology enables one to study corneal morphology layer by layer in extremely thin sections, only disturbing the tissue with an applanating tip. We have demonstrated the cells of the corneal surface, subsurface cells, the topography of Bowman's membrane, corneal lamellae, stromal keratocytes, and the corneal endothelium. The application of this technology lends itself to the in vivo examination of the human cornea. This should aid us greatly in the study of normal morphology, disease states, and the reaction of the cornea in wound healing.

Animals↗
PubMed1993

Confocal microscopy immunofluorescence localization of desmin and other intermediate filament proteins in fetal rat livers.

Immunolocalization of desmin in fetal rat livers shows that on day 12 of gestation a high number of liver cells express desmin. This number decreases from day 14 onward. On day 20 about the same density of desmin-containing cells is found in fetal rat livers as is found in adult rat livers. Desmin-containing cells show two types of labeling patterns, especially on days 12 and 13 of gestation: (a) a basketlike network of intermediate filaments throughout the whole cell, similar to the labeling pattern of cytokeratin in hepatocytes; and (b) more strongly labeled intermediate filaments developing long and slender processes between adjacent cells, close to the labeling pattern of Ito cells in adult rats. From day 14 of gestation, the first type becomes rare, and from day 18 only the second type remains. Double-labeling experiments show that coexpression of desmin and cytokeratin is found in cells of the first type on days 12, 13 and 14 of gestation. Cells containing desmin with the labeling pattern of the second type never express cytokeratin. Coexpression of vimentin and cytokeratin is never found in fetal hepatocytes, even on day 12 of gestation. Numerous nonhepatocyte cells coexpress desmin and vimentin, but some cells contain vimentin or desmin alone. In desmin-containing cells the labeling pattern is of the first type (basketlike network). These results suggest that in early stages of fetal liver development, desmin is found in two different types of liver cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2003

Ligand distributions in agarose particles as determined by confocal Raman spectroscopy and confocal scanning laser microscopy.

Confocal Raman spectroscopy and confocal scanning laser microscopy have been used to analyze ligand distributions within individual chromatographic adsorbent particles. Three different types of particles have been investigated. The first type was synthesized to have a uniform distribution of allyl groups, whereas the two others were designed to have a surface layer of sulphopropyl groups and cores containing allyl groups and dextran, respectively. With confocal Raman spectroscopy it was possible to follow the distribution of both the surface layer and the interior. The distribution of sulphopropyl groups was evaluated with both confocal scanning laser microscopy and confocal Raman spectroscopy, whereas the distributions of allyl groups and dextran were evaluated only with the latter method. The results from the confocal measurements showed the expected result with a uniform distribution of allyl groups in the first type of particle and surface layers of sulphopropyl groups and cores with dextran or allyl groups for the two others.

Adsorption↗
PubMed2006

Visualising fouling of a chromatographic matrix using confocal scanning laser microscopy.

Confocal scanning laser microscopy (CSLM) was used to visualise the spatial location of foulants during the fouling of Q Sepharose FF matrix in finite batch experiments and for examining the subsequent effectiveness of clean-in-place (CIP) treatments in cleaning the heavily fouled beads. Beads were severely fouled with partially clarified E. coli homogenate by contacting the beads with the foulant for contact times of 5 min, 1 or 12 h. The use of two different fluorescent dyes, PicoGreen and Cy5.5, for labelling genomic PicoGreen-labelled dsDNA and protein respectively, allowed the direct observation of the chromatographic beads. The extent of fouling was assessed by measuring the subsequent adsorption of Cy5.5-labelled BSA to the beads. Control studies established that the labelling of BSA did not affect significantly the protein properties. In the control case of contacting the unfouled matrix with Cy5.5-labelled BSA, protein was able to penetrate the entire matrix volume. After fouling, Cy5.5-labelled BSA was unable to penetrate the bead but only to bind near the bead surface where it slowly displaced PicoGreen-conjugated dsDNA, which bound only at the exterior of the beads. Labelled host cell proteins bound throughout the bead interior but considerably less at the core; suggesting that other species might have occupied that space. The gross levels of fouling achieved drastically reduced the binding capacity and maximum Cy5.5-labelled BSA uptake rate. The capacity of the resin was reduced by 2.5-fold when incubated with foulant for up to 1 h. However, when the resin was fouled for a prolonged time of 12 h a further sixfold decrease in capacity was seen. The uptake rate of Cy5.5-labelled BSA decreased with increased fouling time of the resin. Incubating the fouled beads in 1 M NaCl dissociated PicoGreen-labelled dsDNA from the bead exterior within 15 min of incubation but proved ineffective in removing all the foulant protein. Cy5.5-labelled BSA was still unable to bind beyond the outer region of the beads. A harsher CIP treatment of 1 M NaCl dissolved in 1 M NaOH was also ineffective in removing all the foulant protein but did remove PicoGreen-conjugated dsDNA within 15 min of incubation. Cy5.5-labelled BSA was able to bind throughout the bead interior after this more aggressive CIP treatment but at a lower capacity than in the case of fresh beads. The competitive adsorption of BacLight Red-labelled whole cells or cell debris and PicoGreen-conjugated dsDNA was also visualised using CSLM.

Biofilms↗
PubMed1999

Apoptosis and morphology in mouse embryos by confocal laser scanning microscopy.

Confocal laser scanning microscopy combined with a vital stain was used to study apoptosis in organogenesis-stage mouse embryos. Apoptosis has previously been visualized in whole embryos using the vital dyes acridine orange, Nile blue sulfate, and neutral red. In the present study, mouse embryos were harvested on Gestation Day 9 and stained with the vital lysosomal dye LysoTracker Red. Following incubation in the stain, embryos were fixed overnight in 4% paraformaldehyde, dehydrated in a graded methanol series, and cleared in benzyl alcohol/benzyl benzoate. The resulting embryo is almost transparent and retains specific LysoTracker Red staining. To achieve optical sectioning through embryos, it was necessary to use low-power objectives. With this procedure, the entire embryo can be optically sectioned and reconstructed in three dimensions to reveal areas of dye staining. Our results demonstrate specific regions undergoing programmed cell death in normal development and increased LysoTracker staining in embryos exposed to hydroxyurea. This procedure allows for the optical imaging of whole Day 9 ( approximately 22 somites) embryos that were greater than 700 microm thick in the z axis and can be applied to studies involving neural tube formation or other aspects of organogenesis.

Aldehydes↗
PubMed1995

Visualization of diffusion pathways across the stratum corneum of native and in-vitro-reconstructed epidermis by confocal laser scanning microscopy.

Confocal laser scanning microscopy is a technique that permits the direct visualization in unfixed material of diffusion pathways and the cellular distribution of fluorescent markers after topical applications. This approach, in which the tissue specimen is optically sectioned, allows the study of changes in distribution pattern of applied compounds depending on the vehicle, time and depth without the interference of chemical alterations induced by most of the current techniques used for such studies. Using this technique the permeability properties of in-vitro-reconstructed epidermis were compared with those of the native counterpart. The epidermis was reconstructed by culturing human adult keratinocytes at the air-liquid interface either on fibroblast-populated collagen or on de-epidermized dermis. A fluorescent probe--Nile red (NR)--was applied in three different vehicles--polyethylene glycol (PEG) with a molecule mass of 400 (Da), propylene glycol (PG) and dimethyl sulphoxide (DMSO)--which perturb the SC barrier function to different extents. When NR was applied in PEG and PG on native epidermis, the amount of NR penetrating into and through the SC was very low, but was markedly increased when NR was applied in DMSO. Unlike native epidermis, the reconstructed epidermis allowed rapid NR penetration after the application in any of the solvents used. Furthermore, NR applied on reconstructed epidermis, was distributed quite homogeneously between the cellular and the intercellular spaces throughout the SC, suggesting that not only intercellular lipid structures but also the properties of the cornified envelopes differed markedly from those found in native epidermis.(ABSTRACT TRUNCATED AT 250 WORDS)

Cells, Cultured↗
PubMed2006

Modeling and simulation of protein uptake in cation exchanger visualized by confocal laser scanning microscopy.

Confocal laser scanning microscopy (CLSM) has been extensively applied in the area of protein chromatography to investigate the uptake mechanism of protein in adsorbents. However, due to the light attenuation in the deeper layers of a specimen, quantitative analysis using CLSM data is still far from reality. In this work, an attenuation equation for describing the darkening of the CLSM image in the deeper scanning layers was developed. Bovine serum albumin (BSA) adsorption to SP Sepharose FF was performed by batch adsorption and micro-column chromatography on which protein concentration in single absorbents were visualized by CLSM. The parameters in the equation were estimated by fitting it to the fluorescence intensity profiles obtained at adsorption equilibrium, and then the equation was used to simulate the effect caused by the light scattering and absorption. CLSM analysis demonstrated that BSA adsorption to SP Sepharose FF followed the shrinking core pattern and was predicted reasonably well by the pore diffusion model in combination with the attenuation equation. By comparison of the CLSM data with the simulations, it shows that the attenuation equation was useful to demonstrate the validity of an intraparticle mass transport model for the estimation of intraparticle protein concentration profiles.

Adsorption↗
PubMed2005

Protein-labeling effects in confocal laser scanning microscopy.

Confocal laser scanning microscopy (CLSM) is being increasingly used for observing protein uptake in porous chromatography resins. Recent CLSM studies have revealed the possible existence of a nondiffusive protein transport mechanism. Observing protein uptake with CLSM requires labeling the protein with a fluorescent probe. This study examines the effect of the probe identity on the subsequent CLSM adsorption profiles. The adsorption of lysozyme conjugated with different fluorescent probes (Cy5, BODIPY FL, Atto 635, and Atto 520) on SP Sepharose Fast Flow was measured using CLSM and zonal chromatography experiments. Results from zonal chromatography show that the retention time of lysozyme-dye conjugates differ significantly from unlabeled lysozyme. The change in retention of lysozyme upon conjugation with a fluorescent probe is consistent with the difference in net charge between the lysozyme-dye conjugate and unlabeled lysozyme. The adsorption profiles measured by CLSM show significantly different behavior depending upon whether the lysozyme-dye conjugate is retained longer or shorter than the unlabeled lysozyme. These results strongly suggest that the lysozyme concentration overshoot observed in previous CLSM experiments is the result of displacement of weaker binding labeled lysozyme by stronger binding unlabeled lysozyme.

Animals↗
PubMed2001

Morphologic features of melanocytes, pigmented keratinocytes, and melanophages by in vivo confocal scanning laser microscopy.

Confocal scanning laser microscopy (CSLM) represents a novel imaging technique for in vivo microscopic analysis of skin lesions at a level of resolution that allows morphologic analysis of microanatomic structures. We investigated the feasibility of recognizing the cellular constituents of pigmented skin lesions, such as pigmented keratinocytes, melanocytes, and melanophages, by CSLM. Fifteen pigmented lesions (five pigmented seborrheic keratoses, and 10 compound melanocytic nevi) from 15 patients were studied, as well as normal skin. After the clinical lesions were imaged by CSLM, they were biopsied or excised for examination by conventional histology for comparison of the morphologic features. In images obtained by CSLM, pigmented keratinocytes were seen as polygonal cohesive cells with variably bright granular cytoplasm. Melanocytes appeared as bright round, oval, fusiform, or dendritic cells. The architectural growth pattern of melanocytes could be analyzed. Melanocytes were identified by their nested growth pattern as aggregates of bright round to oval structures at the dermoepidermal junction or in the superficial dermis. Melanocytes were also recognizable as single cells along the dermoepidermal junction, usually separated from each other by a variable number of keratinocytes. Melanophages appeared as large bright plump cells with ill-defined cytoplasmic borders, usually located around or near vessels of the superficial dermis. Our results demonstrate that the cellular constituents of pigmented lesions can be recognized by CSLM. This technique sets a new paradigm for noninvasive quasihistologic examination of pigmented lesions in vivo and merits further evaluation for diagnostic use.

Diagnosis, Differential↗