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Results for “Confocal microscopy”

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At least 271 records · Page 15Linked to original sources
PubMed2003

Long-term corneal morphology after PRK by in vivo confocal microscopy.

PURPOSE: To examine human corneal morphology and nerve recovery 5 years after photorefractive keratectomy (PRK). METHODS: Fourteen eyes of 14 patients (ages, 27-53 years) who underwent 6-mm diameter PRK for low to moderate myopia (spherical equivalent [SE] -2.5 to -8.0 D) were examined once 5 years after surgery. Nine healthy individuals served as control subjects. Standard biomicroscopy, manifest refraction, and visual acuity tests were performed. The morphology of the corneas was examined by in vivo confocal microscope. Thicknesses of the epithelium and stroma, as well as the density of corneal opacity (haze) were obtained from digital image analysis of the confocal microscopy through-focusing (CMTF) scans. RESULTS: Confocal microscopy revealed increased reflectivity in the subepithelial extracellular matrix, keratocyte nuclei and processes in all patients. The mean objective haze estimate was 166.7 U (range, 50-390) in control corneas compared with a mean of 225.9 U (range, 125-430, P = 0.15) in the post-PRK corneas. The density of the subbasal nerve fiber bundles in post-PRK corneas (mean, n = 4.2; range, n = 1-7 per field of view) was not significantly lowered from that in control subjects (mean, n = 4.9; range, n = 3-6; P = 0.56). Bowman's layer was undetectable in all post-PRK corneas. Clinically, slit-lamp-observed trace of haze in four corneas correlated positively with the ablation depth (P = 0.016) and the thickness of the haze area (P = 0.006) in the confocal microscope. CONCLUSIONS: In vivo confocal microscopy demonstrates the presence of morphologic alterations even 5 years after PRK. However, these alterations are overcome by cellular and neural recovery and do not seem to interfere with visual performance.

Adult↗
PubMed1993

Comparison of anti-fading agents used in fluorescence microscopy: image analysis and laser confocal microscopy study.

To ascertain the ability of commercial and home-made anti-fading media to reduce the decrease of fluorescein isothiocyanate (FITC) fluorescence, we studied the bleaching characteristics of FITC-stained Reh 6 cells mounted in buffered glycerol and in anti-fading media. We measured the intensity of fluorescence over time with a confocal laser scanning microscope and a standard epifluorescence microscope coupled to an image analysis system. Most of the anti-fading media effectively retard fading but each has drawbacks. Better results were obtained with media containing p-phenylenediamine (solutions in buffered glycerol, Vectashield, Fluorstop). However, Mowiol, Slowfade, n-propyl gallate (20 g/liter) were also effective in retarding fading. Most of them, except Mowiol, reduced fluorescence intensity. We concluded that the choice of anti-fading medium would depend on the desired results: a slower decay of fluorescence despite an initial quenching of fluorescence or a lower retardant effect with no decrease in initial fluorescence intensity. Moreover, the combination of Mowiol with another anti-fading medium may be a useful compromise when a strong retardant effect is required without marked quenching of the initial fluorescence.

Animals↗
PubMed2003

Confocal microscopy for real-time detection of oral cavity neoplasia.

PURPOSE: The goal of this study was to characterize features of normal and neoplastic oral mucosa using reflectance confocal microscopy. EXPERIMENTAL DESIGN: Oral cavity biopsies were acquired from 17 patients at the Head and Neck Clinic of The University of Texas M. D. Anderson Cancer Center who were undergoing surgery for squamous cell carcinoma within the oral cavity. Reflectance confocal images were obtained at multiple image plane depths from biopsies within 6 h of excision. After imaging, biopsies were fixed in 10% formalin and submitted for routine histological examination. Reflectance confocal images were compared with histological images from the same sample to determine which tissue features contribute to image contrast and can be potentially imaged using in vivo confocal microscopy. RESULTS: Confocal images were successfully acquired from 15 biopsy pairs from 17 patients. Depth-related changes in cell diameter and nuclear density were observed at multiple anatomical sites within the oral cavity. In squamous cell carcinomas, densely packed, pleomorphic tumor nuclei could be visualized with distinct differences in nuclear density and morphology distinguishable between confocal images of neoplastic and nonneoplastic oral cavity. Other features of noncancerous and cancerous oral tissue that could be identified in the confocal images included areas of inflammation, fibrosis, muscle fibers, and salivary glands. CONCLUSIONS: Our results support the potential for this tool to play a significant role in the clinical evaluation of oral lesions, real-time identification of tumor margins, and monitoring of response to therapeutic treatment.

Biopsy↗
PubMed1999

Confocal microscopy in posterior polymorphous corneal dystrophy.

PURPOSE: To report the distinguishing characteristics of posterior polymorphous corneal dystrophy (PPMD) using confocal microscopy. MATERIAL AND METHODS: Two consecutive patients with PPMD were prospectively examined using a white-light tandem scanning confocal microscope with a 24x/0.60 contact objective. RESULTS: At the level of Descement's membrane, roundish hyporeflective images were found in 1 patient. In the other patient, hyporeflective bands were detected. In both patients, patchy hyperreflective areas were identified. CONCLUSION: Confocal microscopy may allow diagnosis of PPMD by demonstrating the alterations in Descement's membrane. This technique is especially valuable in cases of endothelial decompensation, where slit-lamp and specular microscopy may fail to demonstrate changes in Descement's membrane.

Aged↗
PubMedDate not supplied

Myocardial changes after spironolactone in spontaneous hypertensive rats. A laser scanning confocal microscopy study.

The present study aims to objectivate by laser scanning confocal microscopy, the cardiac structure of the spontaneously hypertensive rats (SHR) treated with different doses of spironolactone, either alone or in combination with an angiotensin converting enzyme inhibitor or with a calcium channel blocker. Thirty SHRs were divided into six groups and treated during 13 weeks as follows: control, spironolactone (5, 10 and 30 mg/kg/day), spironolactone + enalapril and spironolactone + verapamil. Spironolactone action on the SHR blood pressure (BP) was dose-dependent. The cardiac hypertrophy was affected by the treatment with spironolactone (high dose) or a combination of spironolactone and Enalapril. The myocardial structure was less affected by the spironolactone monotherapy (at all doses) showing hypertrophied cardiac myocytes, focal areas of the reactive fibrosis, inflammatory infiltrate. The treatment with spironolactone in combination with enalapril or verapamil prevented these alterations. In conclusion, the monotherapy with spironolactone had only a limited effect in the preservation of the myocardial structure and in the attenuation of the interstitial fibrosis in SHRs, even after high dose. This action on the myocardium is more efficient when the spironolactone (even in medium dose) was associated with enalapril or verapamil.

Angiotensin-Converting Enzyme Inhibitors↗
PubMedDate not supplied

Use of in vivo confocal microscopy to understand the pathology of accidental ocular irritation.

In vivo confocal microscopy (CM) provides a unique ability to section optically through living, intact tissues and organs to characterize qualitatively and quantitatively pathological changes in 4 dimensions (x, y, and z, and time). It involves the capture of real-time images without the need for excision, fixation and processing. In vivo CM principally has been used for evaluation of eyes in patients and laboratory animals but has potential application to studies of other tissues/organs. In vivo CM is being used in human ophthalmology clinics. It has been used as a research tool for quantitative, in situ measurement of corneal wound contraction, fibroblast migration, corneal endothelial cell migration, corneal epithelial cell size and desquamation following contact lens wear and surgery, and the assessment of corneal surface toxicity following application of commonly used ophthalmic preservatives. In vivo CM allows us to (a) characterize changes to a light microscopic (i.e., cellular) level; (b) quantify changes objectively: (c) conduct studies of injury and repair in the same animal and directly correlate microscopic changes to clinical observations over time as this technique is used in the living animal; and (d) conduct comparative studies in humans. Here we present a brief overview of in vivo CM and how we are using it to provide noninvasive, in situ qualitative and quantitative histopathologic characterization of accidental ocular irritation. Our intent is to provide an awareness of this relatively new methodology and one practical application of its use in research. The goal of our work is to provide objective, quantitative data for use in developing and validating mechanistically based in vitro replacement tests.

Animals↗
PubMedDate not supplied

Applications of laser scanning confocal microscopy to the observation of human oocytes and embryos.

Laser scanning confocal microscopy (LSCM) is a powerful and useful tool in developmental biology. The diverse applications of LSCM in biomedical field has led to advances in the microscopes themselves and the synthesis of novel specific probes for the observation of biological structures and the hypothesis of physiological process. LSCM was used to visualize the cellular actin cortex together with the chromatin in human arrested preimplantation embryos and in unfertilized oocytes obtained by in vitro fertilization. In embryos, we observed a majority of multinucleated blastomeres and some with fragmented nuclei or anucleated. LSCM also showed nuclei displaying the chromatin condensation and fragmentation patterns suggesting the phenomenon of apoptosis. Moreover, in arrested fragmented embryos of poor morphology, unequal sized blastomeres often showed cellular fragments without nuclei, compatible with aspects of apoptotic bodies.

Actins↗
PubMed2005

[Contribution of in vivo confocal microscopy to limbal tumor evaluation].

PURPOSE: To explore tumors of the limbus with a new in vivo confocal microscope and to compare the images to histology results. METHODS: We evaluated three tumors in three patients with the Heidelberg Retina Tomograph II, Rostock Cornea Module. A diagnostic and therapeutic excision with adjunctive cryotherapy was performed for each individual. Confocal microscopy was compared to histopathologic sections. RESULTS: Histology identified two dysplasias and one carcinoma in situ. The main pathological features were visible on our images: cytonuclear atypias, epithelial folds into an inflammatory and vascularized conjunctival stroma, fine vessels perpendicular to the surface, a clear limit with normal epithelium, papillomatous organization, and abnormal keratinization. CONCLUSION: Our preliminary study showed that this type of limbal tumor could be explored using in vivo confocal microscopy. We were not able to determine whether there was a microinvasion. This new method could be a diagnostic aid, especially for atypical lesions and for follow-up because of frequent recurrences. Other studies are necessary to confirm our hypothesis.

Adult↗
PubMed1991

Confocal microscopy of genome exposure in normal, cancer, and reverse-transformed cells.

Genome exposure studies were carried out on malignant CHO-K1 and C6 rat glioma cells and their respective, phenotypically normal counterparts (reverse-transformed CHO-K1, and both reverse-transformed C6 glioma and normal rat fibroblasts). Cells were subjected to the nick-translation technique previously developed to make visible the exposed (i.e., DNase I-sensitive) nuclear DNA, and examined by both epifluorescence and confocal microscopy. The confocal microscopy, by permitting examination of sections throughout the nucleus, made possible clearer identification of the regions of exposed and sequestered DNA in the cells studied. A peripheral shell of exposed DNA with some discontinuities was displayed in the great majority of the cells with normal phenotype, but in none of the cancer cells. Both types of cells displayed regions of exposed DNA in the nuclear interior, particularly surrounding the nucleoli. In accordance with previous theoretical proposals we postulate: the peripheral nuclear shell of exposed DNA contains differentiation-specific genes that include the specific growth-control genes and that are functional in normal cells but not in cancer; the exposed genes surrounding the nucleoli may represent housekeeping genes active in both normal and cancer cells; and the DNase I-resistant DNA in the interior of the nucleus we postulate to consist for the most part of genes specific to alternative differentiation states and to be sequestered and inactive. Previous differences in evaluation of roles of peripheral and internal DNA sensitivity to DNAse I hydrolysis appear to be reconciled by this formulation. Identification of exposed DNA may be useful in cancer diagnosis.

1-Methyl-3-isobutylxanthine↗
PubMed2001

Dual-color 4Pi-confocal microscopy with 3D-resolution in the 100 nm range.

We report the development of simultaneous two-color channel recording in 4Pi-confocal microscopy. A marked increase of spatial resolution over confocal microscopy becomes manifested in 4Pi-confocal three-dimensional (3D) data stacks of dual-labeled objects. The fundamentally improved resolution is verified both with densely labeled fluorescence beads as well as with membrane labeled fixed Escherichia coli. The synergistic combination of dual-color 4Pi-confocal recording with image restoration results in dual-color imaging with a 3D resolution in the 100 nm range.

Cell Membrane↗
PubMed2006

Copulatory organ musculature in Childia (Acoela) as revealed by phalloidin fluorescence and confocal microscopy.

Copulatory organs of eight species of the monophyletic taxon Childia were investigated in detail, using phalloidin fluorescence method and confocal microscopy. Childia species were shown to have one, two or several tubular stylets, conical to cylindrical in shape, composed of few to numerous needles. The musculature varied greatly, from the absence of seminal vesicle to extensively developed seminal vesicles with several additional types of specialized muscles. Ten copulatory organ characters were coded and mapped on the total evidence tree. The data obtained permitted to follow the evolution of the Childia stylet and to demonstrate that the structure of the stylet apparatus is largely consistent with the phylogeny of the group (CI=0.75). Possible function of different muscle specializations was discussed.

Animals↗
PubMed2002

[Observation on healing process of corneal lens after epikeratophakia with confocal microscopy].

OBJECTIVE: To investigate the nerve and cell healing process of cryopreservation tissue lens after epikeratophakia by in vivo confocal microscopy. METHOD: Epithelia, stroma and nerves of tissue lens, interface between lens and recipient and stroma and endothelia of recipient were observed by in vivo confocal microscopy and recorded by video in 24 cases of keratoconus from 3 days to 5 years after epikeratophakia. RESULTS: Epithelial layer of lens: The tissue lens was covered by corneal superficial flat cells with wing cells and basal cells on the periphery at 3 - 4 days postoperatively, but the morphology and arrangement of these cells were irregular with low density. Superficial flat cells took shape completely at one month, and the morphology and density of the basal cells tended to be normal at 6 months postoperatively. Subepithelial nerve plexus were scattered at 18 months and their normalization completed at 2 years postoperatively. Stroma of lens: Keratocytes appeared circular, dot-shaped, rod-shaped or reticular in morphology postoperatively. Normal keratocytes were discovered sporadically on the periphery of the lens at 2 years postoperatively. At 5 years postoperatively the density of keratocytes was lower than normal stromal keratocyte in the center of the lens. Stromal nerves of lens: Nerves could grow into tissue lens at 6 months postoperatively, the quantity of nerves increased at 2 years and was still less than normal quantity at 5 years postoperatively. Stroma and endothelia of recipient: There was no change in the stroma and endothelia of recipient. CONCLUSIONS: There is significant difference among healing process of epithelia, nerves of unvital tissue lens after epikeratophakia, but there is little change in the transparency of tissue lens.

Adolescent↗
PubMed2000

Dual labeling with green fluorescent proteins for confocal microscopy.

We report a dual labeling technique involving two green fluorescent protein (GFP) variants that is compatible with confocal microscopy. Two lasers were used to obtain images of (i) mixed cultures of cells, where one species contained GFPuv and another species contained GFPmut2 or GFPmut3, and (ii) a single species containing both GFPuv and GFPmut2 in the same cell. This method shows promise for monitoring gene expression and as a nondestructive and in situ technique for confocal microscopy of multispecies biofilms.

Biofilms↗
PubMed1999

Changes in axon arrangement in the retinofugal [correction of retinofungal] pathway of mouse embryos: confocal microscopy study using single- and double-dye label.

The changes in quadrant-specific fiber order in the retinofugal pathway of the C57-pigmented mouse aged embryonic day 15 were investigated by using single- (1,1'-dioctadecyl-3,3,3',3'-tetramethyl-indocarbocyanine perchlorate; DiI) and double- (N-4-4-didecylaminostyryl-N-methylpyridinium iodide; 4Di-10ASP in addition to DiI) labeling techniques. At this earliest stage of development, before any fibers arrive at their targets, retinal axons display a distinct quadrant-specific order at the optic stalk close to the eye. This order gradually disappears along the stalk and is virtually lost at the chiasm, as shown in single-label preparations. The double-label preparations, in which the population peaks of fibers from two retinal quadrants are shown simultaneously in an image, show a fiber arrangement at the chiasm that is different from the pattern seen in the single-label preparations. A distinct and consistent preferential distribution of fibers from different retinal quadrants is shown in the chiasm. Before the midline, the central part of the cross section of the chiasm is dominated by dorsal fibers, whereas the rostral and caudal parts of the chiasm are dominated by ventral nasal and ventral temporal fibers, respectively. Moreover, the double-label preparations demonstrate a major reshuffling of fiber position after the fibers cross the midline. Fibers from ventral retina are shifted gradually to a rostral position at the threshold of the optic tract, whereas fibers from dorsal retina are shifted caudally. These changes in fiber position indicate a postmidline location in the chiasm, where fibers are re-sorted in accordance with their origins in the dorsal ventral axis of the retina, and suggest a change in axon response to guidance signals when the fibers cross the midline of the chiasm. These changes in fiber order may also be related to the re-sorting of fibers according to their ages at the postmidline chiasm.

Animals↗