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PubMedDate not supplied

Effect of intravenously injected iodinated lipid emulsion on the liver. An experimental study correlating computed tomography findings with in vivo microscopy and electron microscopy findings.

Iodinated lipid emulsions have been shown to have great potential as site specific contrast media for the liver and spleen. Because of unacceptable adverse reactions none of these emulsions has been adopted for clinical use. In an attempt to find an explanation for these adverse reactions we tested three iodinated lipid emulsions, EOE-13, AG 60.99 and AG 66.18. The following models were used: Computed tomography (CT) of the rabbit liver, in vivo microscopy and electron microscopy of the rat liver. The emulsions contained particles of different sizes and were used in varying doses. We found that the larger the emulsion particles, the more likely they were to be taken up by the Kupffer cells and thereby the higher the opacification of the liver achieved at CT. We also observed changes in the microcirculation of the liver when the emulsions were given in doses required to secure satisfactory opacification of the liver at CT. The main changes were 1) a marked increase in the size of the Kupffer cells, and 2) damage to the sinusoidal endothelium, both contributing to sinusoidal congestion. These changes strongly suggest activation of the macrophages and this in turn probably results in the release of toxic mediators. We suspect that the adverse reactions observed in patients when using iodinated lipid emulsions are due to these toxic mediators.

Animals↗
PubMed1976

Observations of the microcirculatory bed in rat mesocecum using differential interference constrast microscopy in vivo and electron microscopy.

The microvascular bed of the rat mesocecum has been examined in vivo using differential interference (Nomarski) optics and subsequently by electron microscopy. The preferential channel, from terminal arteriole to collecting venule, has been examined. In the terminal arteriolar segment the endothelial layer is covered by a continuous layer of smooth muscle cells which, in turn, are surrounded by adventitia. In the metarteriolar segment the periendothelial cells still resemble smooth muscle cells but the tunica media is discontinuous. In the distal segment periendothelial cells are more scattered and have the appearance of pericytes. There appears to be a continuous transition of the periendothelial cell layer from terminal arteriole to distal segment. Nerve endings were seeen in both the terminal arteriolar and metarteriolar segments. During contraction smooth muscle cells, oriented circumferentially, shorten and thicken. Endothelial cells appear anchored by myoendothelial junctions. Endothelial cells have filaments which show increased banding during vasoconstriction, suggesting that such cells may contract. Capillary offshoots leave the preferential channel, usually at right angles. Smooth muscle cells are oriented to form a sphincter and there are many myoendothelial junctions at the branch point. Within a short distance the capillary branch loses its periendothelial coat.

Animals↗
PubMed1993

Dendritic cell/lymphocyte clustering: morphologic analysis by transmission electron microscopy and distribution of gold-labeled MHC class II antigens by high-resolution scanning electron microscopy.

Dendritic cells (DCs) are potent antigen-presenting cells for a variety of immune responses; however, their mechanism of action has not been established. It is known that DCs can cluster with one another and with other cell types during in vitro immune responses, and clustering may be essential for the activation of resting lymphocytes. In this study, ultrastructural examination of clusters that form during extended culture of enriched rat splenic DCs (approximately 70% DCs) is reported. DCs were readily distinguished from other cell types, which included lymphocytes and macrophages. DCs displayed characteristic veils and/or dendritic processes that intertwined with processes of other cells within the cluster, or extended from the cluster periphery. Occasional DCs contained large vacuoles lined with small vesicles. A paramount feature of DCs is their constitutive expression of high levels of surface major histocompatibility complex class II antigens. The surface distribution of class II antigens on clustering DCs was examined using 10 nm immunogold labeling techniques and high-resolution scanning electron microscopy. DCs were readily distinguished by morphologic criteria, and examination of various surface membrane regions revealed a differential distribution of class II antigens. Gold label was frequently distributed in linear arrays and clusters, suggesting a cytoskeletal role in the recycling/redistribution of Class II antigens. These morphologic findings further an understanding of basic DC biology and their mechanism of action as antigen-presenting cells.

Animals↗
PubMed2005

A vision-based, 3D reconstruction technique for scanning electron microscopy: direct comparison with atomic force microscopy.

High-resolution, detailed 3D reconstructions of biological specimens obtained from scanning electron microscopy stereo-micrographs and proprietary software were compared with Tapping-Mode AFM datasets of the same fields. The reconstruction software implements several original solutions including a neural adaptive point-matching technique, the ability to build an irregular triangulated mesh rather than a regular orthogonal grid, and the ability to re-map one of the original images exactly onto the reconstructed surface. The technique was applied to human nerve tissue to obtain 1,424 x 968-pixel, texture-mapped datasets, which were subsequently compared against 512 x 512-pixel AFM datasets from the same viewfields. Accounting for the inherent differences of the two techniques, direct comparison revealed an excellent visual match. The correspondence was also quantified by calculating the cross-correlation coefficient between corresponding altimetric profiles in SEM and AFM data, which consistently exceeded a figure of 0.9, with a rate of point mismatch in the order of 0.01%. Research is still underway to improve the robustness of the technique when applied to arbitrary images

Humans↗
PubMed1981

Structure and porosity of human cervical enamel studied by polarizing microscopy and transmission electron microscopy.

Dehydration by alcohols and concentrated solutions of highly soluble salts caused refractive changes in enamel which were enhanced in the cervical regions. Electron microscopy showed crystals in cervical enamel to vary from normal orientation, size and habit to marked disorientation and reduced size. Consequent increases in surface area and inter-crystalline volume could account for the exaggerated optical behaviour. The changes in optical properties of enamel occurred only when the osmotic pressures of the imbibing solutions exceeded 500 atm. It is proposed that the internal pore system may take up water freely but exclude large hydrated molecules and ions which thereby create a negative pressure on the water contained in these pores. Once this negative pressure exceeds the tensile strength of the water-pore system, molecules may be torn from their attachment sites, and possibly from each other, so that a sudden evacuation of the pores is accompanied by a simultaneous reduction in refractive index. It is likely that such sudden evacuation would alter the potential of the pore surfaces to react with small molecules and ions subsequently invading the tissue.

Absorption↗
PubMed1995

A comparative study of the application of scanning acoustic microscopy and confocal laser scanning microscopy to the structural assessment of human bones.

The Scanning Acoustic Microscope (SAM) is used to image mechanical structure and to measure the sound velocity in micrometers of in vitro samples from human bone. A similar principle is used in confocal laser scanning microscopy. The same sample is imaged by both methods and the results compared. We concentrated on the SAM, the function of which is described in greater detail. We demonstrate that the confocal scanning principle is applied in both microscopes, although information about structure differs, depending on the method used.

Acoustics↗
PubMed2006

Scanning force microscopy and fluorescence microscopy of microcontact printed antibodies and antibody fragments.

Unlabeled primary immunoglobulin G (IgG) antibodies and its F(ab')2 and Fc fragments were attached to oxygen-plasma-cleaned glass substrates using either microcontact printing (MCP) or physical adsorption during bath application from dilute solutions. Fluorescently labeled secondary IgGs were then bound to surface-immobilized IgG, and the relative surface coverage was determined by measuring the fluorescence intensity. Results indicated that the surface coverage of IgG increased with increasing protein solution concentration for both MCP and bath-applied IgG and that a greater concentration of IgG was transferred to a glass substrate using MCP than during physisorption during bath applications. Scanning force microscopy (SFM) showed that patterned MCP IgG monolayers were 5 nm in height, indicating that IgG molecules lie flat on the substrate. After incubation with a secondary IgG, the overall line thickness increased to around 15 nm, indicating that the secondary IgG was in a more vertical orientation with respect to the substrate. The surface roughness of these MCP patterned IgG bilayers as measured by SFM was observed to increase with increasing surface coverage. Physisorption of IgG to both unmodified patterned polydimethylsiloxane (PDMS) stamps and plasma-cleaned glass substrates was modeled by Langmuir adsorption kinetics yielding IgG binding constants of K(MCP) = 1.7(2) x 10(7) M(-1) and K(bath) = 7.8(7) x 10(5) M(-1), respectively. MCP experiments involving primary F(ab')2 and Fc fragments incubated in fluorescently labeled fragment-specific secondary IgGs were carried out to test for the function and orientation of IgG. Finally, possible origins of MCP stamping defects such as pits, pull outs, droplets, and reverse protein transfer are discussed.

Adsorption↗
PubMed2002

A novel sample holder allowing atomic force microscopy on transmission electron microscopy specimen grids: repetitive, direct correlation between AFM and TEM images.

A novel sample holder that allows atomic force microscopy (AFM) to be performed on transmission electron microscope (TEM) grids is described. Consequently, AFM and TEM images were repeatedly obtained on exactly the same sample area. For both techniques, a thin carbon film was used as the imaging substrate. Although these techniques have been previously used in conjunction, AFM and TEM images on exactly the same area have not been repeatedly obtained for any system. Correlation of AFM and TEM images is useful for work where the three-dimensional topographical information provided by the AFM could be used to better interpret the two-dimensional images provided by the TEM and vice versa. To demonstrate the applicability of such correlation, new results pertaining to a fibrillar collagen system are summarized.

Fibrillar Collagens↗
PubMed2002

Comparison of the axial resolution of practical Nipkow-disk confocal fluorescence microscopy with that of multifocal multiphoton microscopy: theory and experiment.

We compare the axial sectioning capability of multifocal confocal and multifocal multiphoton microscopy in theory and in experiment, with particular emphasis on the background arising from the cross-talk between adjacent imaging channels. We demonstrate that a time-multiplexed non-linear excitation microscope exhibits significantly less background and therefore a superior axial resolution as compared to a multifocal single-photon confocal system. The background becomes irrelevant for thin (< 15 microm) and sparse fluorescent samples, in which case the confocal parallelized system exhibits similar or slightly better sectioning behaviour due to its shorter excitation wavelength. Theoretical and experimental axial responses of practically implemented microscopes are given.

Fluorescence Polarization↗
PubMed1990

Thickness measurements on V79-4 cells: a comparison between laser scanning confocal microscopy and electron microscopy.

A quantitative comparison has been carried out between laser scanning confocal microscopy on living cells and standard electron microscope methods on fixed samples. It was estimated from these measurements that there was about 10-20% reduction in thickness in fixed samples of monolayer V79-4 hamster cells. Precise information on the true thickness of living cells, as irradiated, is required for full interpretation of radiobiological data with poorly penetrating radiations, including ultrasoft X-rays. The confocal microscope allows rapid measurements on unperturbed living samples.

Animals↗
PubMed1978

Sephadex globules placed as landmarks in combined stereo microscopy, scanning and transmission electron microscopy.

Regions of specific interest in tissue blocks were localized in a stereo microscope and landmarked with Sephadex spheres (10-40 micron in diameter). This procedure made it possible to recognize these regions easily and quickly in the scanning electron microscope. When the tissue was later embedded in Epon the spheres facilitated orientation when sectioning. Thus it was possible with great certainty to determine the level at which ultrathin sections should be cut for transmission electron microscopy to attain an exact correspondence between scanning and transmission electron microscopic observations. The procedure is described and an example of its application is shown in a study of experimental hypertensive endothelial changes.

Angiotensin II↗
PubMed1983

A routine flat embedding method for electron microscopy of microorganisms allowing selection and precisely orientated sectioning of single cells by light microscopy.

A simple method is described to embed material in resin, in the form of microscope slides, to observe it with high resolution light microscopy, to select, orient and section it for TEM. This method can be applied to many kinds of material but is particularly useful for the study of rare or tiny plant or animal microorganisms from field or culture. A diamond scriber, translucent hydrosoluble resin release agent, translucent and smooth resin stubs and a longitudinally perforated block-holder for ultramicrotome are the specific tools of this method.

Microbiological Techniques↗
PubMed2002

New modal wave-front sensor: application to adaptive confocal fluorescence microscopy and two-photon excitation fluorescence microscopy.

Confocal and multiphoton microscopes are particularly sensitive to specimen- or system-induced aberrations, which result in decreased resolution and signal-to-noise ratio. The inclusion of an adaptive optics correction system could help overcome this limitation and restore diffraction-limited performance, but such a system requires a suitable method of wave-front measurement. By extending the concept of a modal wave-front sensor previously described by Neil et al. [J. Opt. Soc. Am. A 17, 1098-1107 (2000)], we present a new sensor capable of measuring directly the Zernike aberration modes introduced by a specimen. This modal sensor is particularly suited to applications in three-dimensional microscopy because of its inherent axial selectivity; only those wave fronts originating in the focal region contribute to the measured signal. Four wave-front sensor configurations are presented and their input response is characterized. Sensitivity matrices and axial responses are presented.

Microscopy, Confocal↗
PubMed2001

Field emission scanning electron microscopy and freeze-fracture transmission electron microscopy of mouse cerebellar synaptic contacts.

Samples of albino mice were processed by the cryofracture method for scanning electron microscopy and examined with the field emission scanning electron microscope (FESEM). Freeze-etching direct replicas of mice cerebellar cortex were also studied with the transmission electron microscope (FFTEM), as a complementary technique for obtaining higher resolution, three-dimensional correlative images of cerebellar synaptic contacts. At the granular, Purkinje cells and molecular layers, the cryofracture method for FESEM selectively removed the neuroglial cell investment, facilitating the visualization of the outer and inner surfaces of cerebellar synaptic contacts. In addition, FFTEM showed the real extension of perisynaptic neuroglial investment. The outer surface of mossy fiber rosettes and their digitiform processes were seen at the granular layer, making flat and invaginated synaptic contacts with the granule cell dendrites. At the molecular layer, the longitudinal traject of parallel fibers or nonsynaptic segments and their synaptic varicosities were characterized. These latter established synaptic contacts with Purkinje dendritic spines. Fractured parallel fiber endings showed the SE-I images of clustered spheroidal synaptic vesicles and mitochondria and the surrounding cotton-like appearance of Bergmann glial cell cytoplasm. Climbing fibers showed a characteristic crossing-over bifurcation pattern in the white matter and in the three-layer structure of cerebellar cortex, formation of tendril collaterals in the granular layer, topographical relationship with Purkinje cell soma and retrograde collaterals in the molecular layer. The climbing fiber synaptic relationship with Purkinje dendritic spines was characterized, by means of FFTEM, by the presence of large synaptic endings and aggregation of intramembrane particles at the P and E faces of presynaptic endings, characteristic of excitatory synapses.

Animals↗
PubMed1992

Correlative transmission electron microscopy and high resolution scanning electron microscopy studies on the fine structural organization of the chicken pituitary gland.

The present study employs transmission (TEM) and high-resolution scanning electron microscopy (HR-SEM) to examine the inter- and intra-cellular organization of the pars distalis of the chicken anterior pituitary gland. The overall view of the cryofractured surface of the par distalis illustrates the arrangement of different pituitary cells and tissues in the follicles. Fine structural examination by HR-SEM shows that the membrane of the mitochondria has a similar configuration to that of the rough endoplasmic reticulum. Granule-like structures, on the surface of the mitochondrial membrane, are similar in size to ribosomes. Standard imaging and three-dimensional imaging demonstrated the formation of developing granules inside the Golgi sacs and the release of mature granules from the end of Golgi stacks. The occurrence of granule-granule connections suggests that granules may be released by exocytosis in groups or individually.

Actin Cytoskeleton↗
PubMed1991

High voltage electron microscopy and low voltage scanning electron microscopy of human neoplastic cell culture.

Improved procedures were developed to correlate cell culture data with the images provided by advanced ultrastructural technologies. These procedures were compatible with the two main types of cellular behavior: adherent, spreading (melanomas, rhabdomyosarcomas) and non-adherent in suspension (leukemias). The ultrastructure and function of spreading neoplastic cells primarily depend on surface properties of the attaching substrates. Therefore, the films used for cultured cell whole-mount ultrastructural analysis must have adherence features identical to those of standard cell culture vessels. Improved procedures were developed to produce the polystyrene films of required qualities. These films allowed processing of cells for electron microscopy including chemical fixation, cryo-immobilization, and immunolabelling. Furthermore, these polystyrene films permitted observations of the same cell in the high voltage electron microscope to reveal the internal organization and in the low voltage scanning electron microscope to reveal the surface topography. Neoplastic cells in suspension may dramatically change their ultrastructure as a result of interactions with substrates or other cells. Therefore, immobilization of cellular processes must occur rapidly while cells remain in suspension. These processes were cryo-immobilized by high pressure freezing through the use of the newly designed specimen carrier. Procedures allowing high yield attachment of cryo-fixed neoplastic cells to amino-propyl-derived glass carriers enabled observations of cell surface topography. Furthermore, freeze-substitution and drying of freeze-fractured cells revealed their three-dimensional internal organization in the low voltage scanning electron microscope.

Cryopreservation↗
PubMed1990

[Light microscopy and electron microscopy study of the lymphatic capillaries of human dental pulp].

The existence and the morphological characteristics of the lymphatic capillaries of the dental pulp have been and are yet discussed. In this study the morphological properties of the lymphatic capillaries of the healthy human dental pulp have been described at light and electron microscopy. For the investigation human enclosed incisive and canine teeth surgically extracted and permanent premolars extracted for orthodontic reasons were used. On semithin sections the lymph capillaries are detectable as vessels of 15-50 microns in diameter with a very irregular shape and a subtle wall. No pericytes or muscular cells surround the endothelial lining. At ultrastructural level the lymph capillaries show a very thin and indented wall with protrusions towards the vessels lumen or the interstitium. The connections between adjacent endothelial cells are different and variously shaped: end to end contacts, overlapping between cellular protrusions and complex interdigitations among multiple protrusions of the endothelial cells. Sometimes the overlappings between endothelial cells determine intraparietal channels in communications with the interstitium and the vessel lumen. A discontinuous basal lamina and a network of filaments and fibrils surround the endothelial wall. Between the cytoplasmic organelles micropinocytotic vesicles, multivesicular bodies and the Weibel-Palade bodies are present.

Basement Membrane↗