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At least 1,693 records · Page 94Linked to original sources
PubMed1998

A simplified procedure for the physical development of the sulphide silver method to reveal synaptic zinc in combination with immunocytochemistry at light and electron microscopy.

The pool of zinc present in excitatory synaptic terminals in normal and pathological conditions (for instance the status epilepticus induced by kainic acid) can be stained by a silver sulphide method followed by physical development of the insoluble zinc-sulphide complexes. In this study we applied a previously described simple and rapid developing procedure that reveals synaptic zinc, to the study of normal and pathological hippocampi and combined it with pre and postembedding immunocytochemical methods to detect different antigens. Normal and kainic acid-treated rats were perfused with fixative solutions containing sodium sulphide and 50 microm-thick vibratome sections of the hippocampi were incubated in a commercial developing solution (IntenSE M, Amersham). The developed vibratome sections were then (1) mounted for light microscopy or osmicated and epon-embedded for electron microscopy; or (2) processed for the preembedding immunoenzymatic detection of various antigens (GABA, parvalbumin, calbindin) with light and electron microscopy. Thin sections from epon-embedded samples were also processed for the postembedding immunogold localization of glutamate. This very simple and rapid procedure gives rise to zinc-specific staining, comparable to that obtained with classical developing methods and good preservation of both antigenicity and ultrastructure. It is therefore possible to detect, in the same thick or thin section, zinc reaction product and different antigens.

Animals↗
PubMed1998

Detection of viruses by electron microscopy: an efficient approach.

Wetting of grids is the most important aspect of the detection of viruses by electron microscopy. Carbon or plastic/carbon coated grids used in most electron microscopy laboratories are negatively charged and hydrophobic (Sogo, Portman, Kaufman and Koller, 1975. J. Microsc. 104, 187-198.). Most viruses are negatively charged at pH 6.0-7.0 which does not allow them to adhere effectively to the support film. Viruses are then washed away or form clumps on the grid making detection more difficult. A suitable wetting agent, 0.1% poly-L-lysine hydrobromide (MW 35000) has been used for the detection of plant viruses (Roberts, personal communication). This chemical was used for processing faecal samples from patients with gastroenteritis for electron microscopy. The result showed a significant increase in terms of virus concentration on the grids and an increase in the general detection rate of positive samples. A study was then undertaken comparing the treatment method of grids using faecal samples. The grids were treated and not treated with poly-L-lysine hydrobromide.

Adenovirus Infections, Human↗
PubMed1985

Present position of microscopy and of culture in diagnostic mycobacteriology.

The relative merits of microscopy and culture are discussed; it is attempted to review the possible reasons why microscopy is considered as unreliable in TB diagnosis, including the probably false application of Bayes' theorem. In the author's opinion, microscopy still holds a firm position in the rapid and relatively reliable detection of pulmonary TB. As for culture, it is highly questionable if "better media" could accelerate the growth rate of tubercle bacilli. Therefore, it should be attempted to introduce novel techniques which will allow to decrease considerably the mycobacterial mass needed for detection. Laboratories equipped with such devices could offer their services even on an intercontinental basis, in order to have a favourable cost-benefit ratio.

Bacteriological Techniques↗
PubMed2005

[In vivo confocal microscopy and ocular surface diseases: anatomical-clinical correlations].

PURPOSE: To describe corneal and conjunctival epithelial changes in various ocular surface disorders with a new in vivo confocal microscope (Corneal HRT II module) and to compare these results with those obtained by immunohistology on impression cytology specimens. METHODS: In this study, we investigated 36 eyes of 18 patients with ocular surface diseases such as corneal epitheliopathy induced by ocular rosacea (five patients), keratoconjunctivitis sicca related to tear evaporative condition in Meibomian gland dysfunction (eight patients), or tear-deficient dry eye in Sjögren's syndrome (five patients). In vivo confocal microscopy imaging (Heidelberg Retina Tomograph II Rostock Cornea Module) and impression cytology of the cornea and/or conjunctiva were performed. The images were analyzed for epithelial cell morphology at nuclear and cytoplasmic levels, in the cornea and the conjunctiva, number of goblet cells and their possible presence within the corneal epithelium, corneoconjunctival junction at the limbus, and density and presence of inflammatory cells within both epithelia. RESULTS: Images obtained consisted of two-dimensional 400x400 microm optical sections oriented parallel to the surface of the eye and compared with impression cytology. Conjunctival and corneal epithelia showed dramatic changes in ocular surface disease. In keratoconjunctivitis sicca, we found squamous metaplasia, inflammatory cell infiltration, goblet cell depletion, as well as a nuclear snake-like chromatin pattern. In rosacea associated with corneal epitheliopathy, a corneal conjunctivalization (goblet cells together with conjunctival epithelial cells within the corneal epithelium layer) was found. The images provided by this technique are in excellent correlation with the findings using impression cytology. CONCLUSION: The HRT II in vivo confocal microscope facilitates the study of both central and peripheral ocular surface epithelia compared to first-generation confocal microscopy devices. In vivo confocal microscopy, showing corneal and conjunctival epithelial structures in the live eye at the cellular level, is likely to open up a new promising way to investigate ocular surface involvement in complex diseases, providing a new insight on corneal and conjunctival disorders in the future.

Conjunctiva↗
PubMed1989

Spontaneous lesions in the respiratory epithelium of the Syrian golden hamster as seen by scanning electron microscopy.

The respiratory epithelium of 8, 12.5 and 17.5-month-old Syrian golden hamsters was investigated by scanning electron microscopy (SEM) and routine light microscopy (LM) of paraplast sections. In selected cases, SEM-specimens were embedded in Epon after SEM evaluation. The samples were cut in semithin sections and examined by light microscopy. The density of ciliated cells in the respiratory epithelium differs from hamster to hamster. Care must, therefore, be taken when diagnosing simple metaplasia by SEM alone unless a sufficient number of damaged cilia are present. Only 1 animal (8 months old) exhibited squamous metaplasia of the tracheal mucosa. However, surface polymorphism resembling squamous metaplasia was seen in almost every specimen. The polymorphism was caused by either submucosal calcifications or by cystic changes of hypertrophic submucosal glands. In addition, variously formed aggregations of mucus were seen protruding from duct openings of hypertrophic submucosal glands. To avoid false positive or negative diagnosis which can occur when screening is done by SEM alone, light-microscopical examination of sections cut from SEM identified lesions appears necessary.

Animals↗
PubMed2000

A new approach for glomerular lesions: evaluation of scanning acoustic microscopy (SAM) for experimental glomerular disease in rats.

Most pathological evaluations using ordinary optical microscopy are usually qualitative and subjective. The beneficial properties of scanning acoustic microscopy (SAM) include not only observation of microstructure but also quantitative measurement of acoustic propagation speed, indicating the tissue elasticity. In this study, we examined the capability of SAM to evaluate pathological findings in glomeruli using anti-Thy.1 glomerulonephritis. Light microscopic observations of the glomeruli showed severe cell proliferation and mesangial matrix expansion at 10 days after induction of glomerulonephritis and, yet, to a lower extent at day 21 with onset of healing. C-mode scanning of SAM enabled imaging of glomerular structure compatible to findings of ordinary light microscopy. In addition, glomerular propagation speed in nephritic rats was significantly increased at day 10, and then decreased at day 21. These results indicate that SAM evaluation may be a new, useful technique for quantitative evaluation of proliferative glomerular lesions.

Animals↗
PubMed2002

Detection of Barrett's epithelium by acoustic microscopy.

Barrett's esophagus is associated with increased risk of adenocarcinoma of the gastroesophageal junctional region. The presence of goblet cells (intestinal metaplasia) in columnar cell-lined esophageal mucosa defines Barrett's change. The diagnosis of Barrett's esophagus is based on the presence of intestinal metaplasia in a biopsy from an endoscopically visualized abnormal columnar epithelium. In this pilot study, acoustic microscopy was used to identify the mucosal structure of 10 distal esophageal biopsies. Sections cut at 5 microm of archival paraffin blocks on glass slides were used for this study. Acoustic microscopy permitted the identification of low- and high-power images of epithelial architecture and cellular detail, including Barrett's epithelium. This modality of visualization has the potential to detect lesions such as Barrett's metaplasia, low- and high-grade dysplasia and early carcinoma. If it can be applied to in vivo endoscopy, acoustic microscopy has the potential to increase the accuracy of the diagnosis of Barrett's esophagus, dysplasia and malignancy by providing a method of accurately directing biopsies at endoscopy.

Barrett Esophagus↗
PubMed1978

The microcirculation in two transplantable melanomas of the hamster. II. Scanning electron microscopy.

Normal and tumor vessels observed in vivo on the hamster cheek pouch membrane in transparent chambers were examined by scanning electron microscopy. Large, thin walled veins were found in the reactive zone around the 2 amelanotic melanomas of the hamster. Sinusoidal vessels and channels contained red cells, which did not possess any vascular wall, were seen in the main substance of the tumors. In this system all the vessels within the central mass of the tumor transplant comprise newly formed vessels. The surfaces of the tumor cells and the lining of the tumor vessels were examined by this technique. The biological history i.e., age of the vessel and location from the growing edge of the tumor, were known. The surface of the tumor cells on cut section differed in appearance from the areolar tissue of the pouch membrane. Scanning electron microscopy of the inner aspect of newly formed tumor vessels permits much greater sampling of the tissue than transmission electron microscopy and further work should reveal aspects of the interaction of the blood with tumor stroma.

Animals↗
PubMed2001

Scaling-index method as an image processing tool in scanning-probe microscopy.

The scaling-index method (SIM) is a novel tool for image processing in scanning-probe microscopy. Originating from the theory of complex systems, the SIM can be used in order to extract structural information from arbitrary data sets. This method can readily be applied to the analysis of digital atomic-force microscopy (AFM) images. Especially for biomedical diagnostics, where genetic material is investigated by various microscopic methods, a reliable image segmentation based on the SIM algorithm is helpful. As a first application, AFM-images of GTG-banded human metaphase chromosomes (with G bands obtained by Trypsin using Giemsa) are compared with micrographs from conventional light microscopy by means of a scaling-index analysis. While the grey-level distributions of the optical and the AFM-images are largely different from each other, the scaling-index images are remarkably similar. Using this method, a fingerprint of an image can be produced which helps in the classification and interpretation of the measured data.

Algorithms↗
PubMed1999

The role of persisting infections in the pathogenesis of pulmonary emphysema. Electron microscopy reveals a probable bacterial colonization of the alveolar space and the bronchioles.

Lung volume reduction surgery (LVRS) yields resection specimens from patients with advanced pulmonary emphysema. Regarding the development of lung function parameters, recent results obtained by light microscopy revealed an unfavorable prognosis in patients with remarkable inflammation, particularly in the bronchioli. Tissue from ten patients (alpha1-antitrypsin level in the normal range) was furthermore investigated by electron microscopy. Scanning electron microscopy shows 0.4-0.6 micron spherical bodies variably densely arranged in the whole alveolar space and in the bronchioles of all patients. These bodies are mostly seen on the microvilli of type II pneumocytes. An immunological reaction with activation of macrophages and granulocytes occurs simultaneously. Macrophages show cytoplasmic extensions to the spherical bodies, which exhibit a cellular membrane but no cellular wall. This favors the diagnosis of bacterial colonization of the alveolar space and the bronchioles by mycoplasmas or L-forms of other bacteria. As patients undergoing lung volume reduction surgery are under optimal medical treatment and without any infection clinically, these findings appear to be relevant for the pathogenesis and/or progression of pulmonary emphysema.

Acute Disease↗
PubMed1996

Detection of 'rare event' fetal erythroblasts in maternal blood using automated microscopy.

This paper describes the use of automated microscopy to detect fetal erythroblasts in maternal blood. The technology is based on the following approach: (1) the use of centrifugal cytology for the preparation of monolayers; (2) simultaneous staining of fetal hemoglobin (immunoalkaline phosphatase) and chromosome sequences (FISH); (3) multi-mode microscopy to detect rare events; (4) visual evaluation of image memories containing detected objects. Model systems show that fetal cells in frequencies as low as 1 in a million cells can be detected easily (manually or by automated microscopy). Algorithms for automated cell selection were developed for a test set of 6 patients. Optimization of hardware and software routines will make analysis of several million cells in approximately 1 h feasible.

Autoanalysis↗
PubMed1999

Transdermal water mobility in the presence of electrical fields using MR microscopy.

Magnetic resonance microscopy of skin from hairless rats under the influence of electrical fields was conducted for two cases: 1) low voltage constant electrical fields and 2) high-voltage short pulse, electrical fields. Under conditions of the low voltage and low current iontophoresis, i.e., 0 to 20 V, and 0 to 0.5 mA/cm2, it was found that the skin structure, as observed by magnetic resonance microscopy, did not significantly change until 20 Volts were applied across the 0.1 cm thick skin. Under these conditions, the viable epidermis appeared to swell, and this result corresponded to observations from scanning electron microscopy and other research from the literature. High voltage electrical fields, i.e., 220 V 1 ms pulses repeated once per second, appeared to hydrate the stratum corneum as is consistent with published literature on electroporation. In the case of iontophoresis, water self-diffusion coefficients in the epidermis and hair follicle regions at all voltages were affected by the electrical field. Statistical analysis at the 95% confidence level for the comparison of the average differences between diffusion coefficients with the electrical field on and with the electrical field off for pair matched pixels for the viable epidermis show that for 5 V (p = 0.00377), 10 V (p = 0.0108), 20 V (p = 0.0219) regimes there are statistically significant (p < or = 0.05) changes due to the applied electric field. The same analysis for the hair follicle region at 5 V (p = 6.89 x 10(-7)), 10 V (p = 1.42 x 10(-5)), 20 V (p = 3.23 x 10(-3)) also show statistically significant changes (p < or = 0.05). When the electroporation pulse was applied, the water diffusion coefficients increased by about 30% to 6.6 x 10(-6) cm2/s +/- 2.4 x 10(-7) cm2/s and 8.3 x 10(-6) cm2/s +/- 3.7 x 10(-7) cm2/s, for the epidermis and hair follicle regions, respectively. Significant differences were noted between diffusion coefficients in the viable epidermis and the hair follicles for all cases.

Animals↗
PubMed1999

Atomic force microscopy imaging of dried or living bacteria.

Atomic force microscopy (AFM) was used to obtain micrographs of dried bacteria in air, and of living ones in their culture medium. Images of dried bacteria were very similar to images obtained elsewhere by the much more complicated cryoetching preparation technique for transmission electron microscopy. Living bacteria were immobilized on a poly-L-lysine film, and directly observed in their culture medium at a resolution unattainable by any other technique applicable to living material. The images were similar to those obtained in scanning electron microscopy where the specimen must be fixed, dried and coated with conductive material, and as a result, no longer viable.

Desiccation↗
PubMed1998

Electron microscopy study of explanted intraocular lenses from clinically noninfected eyes.

PURPOSE: To determine whether intraocular lenses (IOLs) in clinically noninfected eyes are coated with a significant, bacteria-containing biofilm. SETTING: The Oxford Eye Hospital, Oxford, United Kingdom. METHODS: Twenty-six IOLs, removed for reasons other than endophthalmitis from 26 patients attending the Oxford Eye Hospital over a 3 year period, were examined by electron microscopy. Immediately following explantation, the IOL was placed in glutaraldehyde 4% in 0.1 M phosphate buffer solution and processed for scanning electron microscopy (SEM). Areas of interest were reprocessed for transmission electron microscopy (TEM). RESULTS: There was no evidence of a bacterial biofilm on any IOL. In 5 IOLs, significant host cellular debris was seen at the tip of the haptic or at the optic-haptic junction. In 4 of them, clusters of coccoid-shaped structures were seen at the optic-haptic junction on SEM, but examination by TEM showed these structures to be melanosomes, not bacteria. CONCLUSION: We found no evidence to suggest that a significant number of IOLs are coated with a bacterial biofilm in clinically noninfected cases. We advocate the use of TEM to distinguish between coccoid bacteria and melanosomes.

Adult↗
PubMed2000

Optical and atomic force microscopy of an explanted AcrySof intraocular lens with glistenings.

PURPOSE: To assess the surface morphology and cause of glistenings in an explanted AcrySof intraocular lens (IOL). SETTING: Shakai Hoken Kobe Central Hospital, Kobe, Japan. METHODS: A 63-year-old Japanese man had implantation of an AcrySof IOL in the capsular bag. One month postoperatively, he had a neodymium:YAG laser capsulotomy for posterior capsule opacification, which changed the IOL's position in the capsular bag. A few months later, the patient developed disabling night glare from intralenticular glistenings and progressive hyperopic refractive error. The IOL was explanted and then analyzed by optical microscopy and atomic force microscopy (AFM). Laboratory analysis of control AcrySof IOLs kept in a balanced salt solution at steady room and body temperature for 2 months was also performed to evaluate the cause of the glistenings observed clinically. RESULTS: Optical microscopy showed that the explanted AcrySof IOL had several microvacuoles; no abnormalities were observed in the control AcrySof IOLs before or after folding at the room and body temperatures. The AFM analysis showed a significant change in the surface morphology of the explanted IOL, including vacuolar formations in the posterior surface as well as numerous anterior surface irregularities. No microvacuoles or surface morphology alterations were observed in the control AcrySof IOLs by AFM analysis. CONCLUSIONS: The glistenings in the explanted AcrySof IOL were likely caused by temperature changes and not mechanical stress from folding.

Acrylic Resins↗
PubMed1993

Effect of growth of Bacteroides fragilis at different redox levels on potential pathogenicity in a HeLa cell system: demonstration by confocal laser scanning microscopy.

During trauma, the intestinal anaerobe, Bacteroides fragilis, may enter into a pathogenic state. The process coincides with changing environmental conditions particularly the redox level in situ. To gain insight into this phenomenon B. fragilis was grown at different redox levels, and the invasive potential was examined using an in vitro model consisting of HeLa cell monolayers. The clinical strain AIP 5-86 was taken from a small collection of B. fragilis strains able to penetrate into tissue cell monolayers when selected by an acridine orange-crystal violet fluorescent staining technique. Following preliminary investigation by confocal laser scanning microscopy (CLSM), this particular strain was regarded as representative for examining the invasive potential. After growth in a defined medium under oxidizing, reducing or intermediate Eh7 conditions, the washed mid-log phase bacteria were allowed to interact with HeLa cell monolayers for 45 min at 37 degrees C. The results were extensively monitored by CLSM to follow the reactions in a stereoscopic dimension. In addition, the bacteria were examined by transmission and scanning electron microscopy before interaction to distinguish characteristics in surface configuration. The growth of the bacteria at particular redox levels seemed to influence their potential for pathogenicity. After growth at relatively high Eh, the bacteria easily penetrated into the HeLa cells, but not at low Eh, as determined by the laser scanning studies. Examination of the bacteria alone by transmission and scanning electron microscopy revealed small vesicles and a tendency to aggregate after growth at the low redox level while there were rather few vesicles and an implied dispersion at the high redox level. This leaves it open whether the invasiveness was based on the alterations found during growth of the bacteria. Different redox levels as well as the respective changes of the bacterial surface may help to discern the commensal from the pathogenic state of B. fragilis.

Bacteroides fragilis↗
PubMed2003

Crystallisation of estradiol containing TDDS determined by isothermal microcalorimetry, X-ray diffraction, and optical microscopy.

Transdermal drug delivery systems (TDDS) enable a controlled delivery of drugs to the skin. However, it is still a problem to achieve a stable and prolonged constant drug release. To attain high permeation rates across the skin, the concentrations of the drug dissolved have to be high and often create supersaturated, thermodynamically metastable, or unstable systems that possess a high tendency to crystallise. In the present study, microcalorimetry as well as polarisation microscopy and X-ray powder diffraction (XRPD) were used to characterise the growing crystal germs of estradiol (E2) hemihydrate. Polarisation microscopy enabled the observation of crystals with two different morphologies of E2 in the polymeric acrylic transdermal patch matrix. Crystal formation and growth were also detected by XRPD. The diffraction pattern corresponded to estradiol hemihydrate. The intensity of the observed reflections was proportional to the crystal quantities and increased during storage. A high supersaturation resulted in high peak intensities caused by a high crystallisation rate. Since precipitation is generally accompanied by heat evolution, crystal germ formation, and crystal growth could easily be detected early by isothermal microcalorimetry. Much lower amounts of crystal were detected by this method than with the significantly less sensitive XRPD method. Microscopy was equally sensitive to but much more time-consuming than microcalorimetry.

Administration, Cutaneous↗
PubMed2004

Determination of the physical state of norethindrone acetate containing transdermal drug delivery systems by isothermal microcalorimetry, X-ray diffraction, and optical microscopy.

Transdermal drug delivery systems (TDDS) enable a controlled drug delivery to the skin. The low permeability of the stratum corneum necessitates a high drug concentration of the polymeric matrix and often requires supersaturation. This, however, promotes crystallisation of supersaturated systems. Isothermal microcalorimetry at 25 degrees C, polarisation light microscopy, and X-ray powder diffraction (XRPD) were used to characterise the crystal growth of norethindrone acetate (NEA). The solubility of NEA in the patches determined by these methods is about 4%. The crystallisation process could be measured reliably and with a high accuracy by microcalorimetry and microscopy. XRPD was considerably less sensitive but was the only method allowing a semi-quantitative determination of the amounts of crystals formed. The drug-associated heat measured by microcalorimetry increased proportionally with increasing NEA concentration in the concentration range of 4-10% demonstrating a constant crystallisation rate. At a higher supersaturation, such as 12% drug content, the crystallisation process was accelerated. The application of Johnson-Mehl-Avrami kinetics for the analysis of nucleation and crystal growth of the punched patches indicated a site-saturated nucleation mechanism and a one-dimensional crystal growth. The crystallisation enthalpy of NEA was -22.8+/-2.6 kJ/mol. The most specific method to observe the crystal formation is polarisation light microscopy. However, the microscopic analysis requires much longer storage times than microcalorimetry to detect crystallisation.

Administration, Cutaneous↗