Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Microscopy, Interference”

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 865 records · Page 48Linked to original sources
PubMed1992

Costameres are sites of force transmission to the substratum in adult rat cardiomyocytes.

Costameres, the vinculin-rich, sub-membranous transverse ribs found in many skeletal and cardiac muscle cells (Pardo, J. V., J. D. Siciliano, and S. W. Craig. 1983. Proc. Natl. Acad. Sci. USA. 80:363-367.) are thought to anchor the Z-lines of the myofibrils to the sarcolemma. In addition, it has been postulated that costameres provide mechanical linkage between the cells' internal contractile machinery and the extracellular matrix, but direct evidence for this supposition has been lacking. By combining the flexible silicone rubber substratum technique (Harris, A. K., P. Wild, and D. Stopak. 1980. Science (Wash. DC). 208:177-179.) with the microinjection of fluorescently labeled vinculin and alpha-actinin, we have been able to correlate the distribution of costameres in adult rat cardiac myocytes with the pattern of forces these cells exert on the flexible substratum. In addition, we used interference reflection microscopy to identify areas of the cells which are in close contact to the underlying substratum. Our results indicate that, in older cell cultures, costameres can transmit forces to the extracellular environment. We base this conclusion on the following observations: (a) adult rat heart cells, cultured on the silicone rubber substratum for 8 or more days, produce pleat-like wrinkles during contraction, which diminish or disappear during relaxation; (b) the pleat-like wrinkles form between adjacent alpha-actinin-positive Z-lines; (c) the presence of pleat-like wrinkles is always associated with a periodic, "costameric" distribution of vinculin in the areas where the pleats form; and (d) a banded or periodic pattern of dark gray or close contacts (as determined by interference reflection microscopy) has been observed in many cells which have been in culture for eight or more days, and these close contacts contain vinculin. A surprising finding is that vinculin can be found in a costameric pattern in cells which are contracting, but not producing pleat-like wrinkles in the substratum. This suggests that additional proteins or posttranslational modifications of known costamere proteins are necessary to form a continuous linkage between the myofibrils and the extracellular matrix. These results confirm the hypothesis that costameres mechanically link the myofibrils to the extracellular matrix. We put forth the hypothesis that costameres are composite structures, made up of many protein components; some of these components function primarily to anchor myofibrils to the sarcolemma, while others form transmembrane linkages to the extracellular matrix.

Actins↗
PubMed1982

The area of attachment of cytotoxic T lymphocytes to their target cells shows high motility and polarization of actin, but not myosin.

Conjugates of cytotoxic T lymphocytes attached to their target cells were studied by double immunofluorescence on fixed smears to detect simultaneously the localization of actin and myosin within the cells. Actin was found to be polarized in the area of attachment of the lymphocytes (but not of the target cells), whereas myosin remained evenly distributed in the cytoplasm. When living conjugates were brought to 37 degrees C to induce cytotoxicity, this pattern remained unchanged, but observation by interference reflexion microcinematography revealed a high motility of the lymphocytes in the contact area. This localized motility in the area of attachment associated with a peculiar actin polarization, which has no equivalent in any type of cell contact presently known, could represent a necessary step in the sequence of events leading to target cell killing by cytotoxic T lymphocytes.

Actins↗
PubMed2002

Differential interference contrast x-ray microscopy with twin zone plates.

X-ray imaging in differential interference contrast (DIC) with submicrometer optical resolution was performed by using a twin zone plate (TZP) setup generating focal spots closely spaced within the TZP spatial resolution of 160 nm. Optical path differences introduced by the sample are recorded by a CCD camera in a standard full-field imaging and by an aperture photodiode in a standard scanning transmission x-ray microscope. Applying this x-ray DIC technique, we demonstrate for both the full-field imaging and scanning x-ray microscope methods a drastic increase in image contrast (approximately 20x) for a low-absorbing specimen, similar to the Nomarski DIC method for visible-light microscopy.

Journal Article↗
PubMed2000

Computer-controlled impalement of cells in retinal wholemounts visualized by infrared CCD imaging on an inverted microscope.

We present a computer-guided microelectrode positioning system that is routinely used in our laboratory for intracellular electrophysiology and functional staining of retinal neurons. Wholemount preparations of isolated retina are kept in a superfusion chamber on the stage of an inverted microscope. Cells and layers of the retina are visualized by Nomarski interference contrast using infrared light in combination with a CCD camera system. After five-point calibration has been performed the electrode can be guided to any point inside the calibrated volume without moving the retina. Electrode deviations from target cells can be corrected by the software further improving the precision of this system. The good visibility of cells avoids prelabeling with fluorescent dyes and makes it possible to work under completely dark adapted conditions.

Animals↗
PubMed1980

Formation of cell-to-substrate contacts during fibroblast motility: an interference-reflexion study.

The formatin of close contacts and focal contacts (ca. 30 nm and 10-15 nm separation distance respectively) has been studied during the movement of chick heart fibroblasts on planar substrates using interference-reflexion microscopy, and evaluated in the context of spreading and net movement. During spreading the overall advance of the margin of the close contact is steady, punctuated by periods in which it remains stationary, and only 5% of the time is spent withdrawing in contrast to the extreme leading edge. The close contact advances only where a lamellipodium has first extended free of the substrate (greater than or equal to 100 nm separation distance) ahead of the existing close contact. The new close contact is formed by the lamellipodium lowering to the substrate either progressively from its base forward or distally in patches which later join with the main close contact. New focal contacts are formed successively ahead of existing ones, either by microspikes or lamellipodia contacting the substrate locally ahead of the close contact, or within the close contact usually immediately, but not more than 1-2 microns, behind its margin. Examining the cell margin alternately with interference-reflexion and differential-interference contrast showed that the formation of the focal contact was preceded in 90% of the cases by the development of a linear structure in the form of a microspike (as expected), a short projection (< 2 microns long) of the lamellipodium, or a fibre within the lamellipodium, each of which could be traced to the cytoplasmic fibre typically associated with the focal contact. Stress fibres subsequently developed centripetally from these initial fibres. The different forms of the linear structure which preceded the focal contact were interchangeable, giving rise to one another, and we have evaluated that the structure common to each is probably a short bundle of microfilaments. The following features indicate that the close contact plays a primary role in marginal spreading: it is lost when spreading ceases; it is reformed when spreading resumes but only under the newly spread area; the advance of the margin of the thicker leading lamella closely follows that of the close contact; the advance of both can occur ahead of and is thus independent of existing focal contacts and associated stress fibres. We propose that the close contact provides the adhesion required to transmit to the substrate the forces involved in the forward movements of the marginal cytoplasm. The continual formation of focal contacts and stress fibres at the margin is consistent with their role, suggested by others, in drawing the bulk of the cell forward. These evaluations are discussed in the context of the form and distribution of contractile proteins in the cell margin. A primary role of the lamellipodia and microspikes in extending the cell margin and forming new adhesions, preparatory to further cytoplasmic movement, is established by this work.

Animals↗
PubMed1996

[Adhesions and interference images of unfixed erythrocytes].

For the analysis of deformability in microcirculatory investigations an exact understanding of red blood cell (RBC) geometry is required. To extend knowledge we introduce a new morphological feature of resting unfixed erythrocytes by means of an inverted reflection contrast microscope (RCM). By assessing the interference patterns caused by RCM erythrocytes can be classified according to the depth of their central concavity which depends on the flexibility of the RBC. Moreover, the RBC adhesion can be directly observed. We found out that: 1. Five types of normocytes can be distinguished in RCM. 2. In phase contrast the size distribution of RBC without central concavity (type 5, 11.53% of all normocytes) shows peaks at 48 microns2 and 52 microns2. 3. Image analysis reveals two size categories of relative adhesion areas. One category consists of type 1 and 2 (relative adhesion area 25.63%), the other of the types 3 to 5 (relative adhesion area 39.91%). Besides, RCM allows the reliable identification of pathologic erythrocytes in unstained specimens.

Cell Adhesion↗
PubMed1978

Histophysical and histochemical investigations on muscle arterioles in hypertensive arteriolosclerosis; an interference microscopical, polarization optical and histophotometrical study.

In 50 formol fixed biopsies from the m. quadriceps femoris normal and hypertensive arteriolosclerotic arterioles were characterized by quantitative histophysical and histochemical methods. The interference microscopical measurements showed a real increase of dry mass concentration in the pathologically changed vessels. Additional histophotometrical results are interpreted as an increase of proteins with higher isoelectric point (globulins, actomyosin). An augmentation of actomyosin is compatible with the medial hyperplasia demonstrated histologically. Likewise a positive correlation was found between the fibrin incorporation proved histochemically and the quantitatively evaluated protein reaction with Fast green; Glycosaminglycans do not carry weight quantitatively. The polarization optical measurements yielded an alteration of the submicroscopical structure in arteriolosclerosis. It consists in a diminution of the submicroscopical volum of gaps and in a diminished orientated dye binding of Congo red. Thereby the pathogenesis of hypertensive arteriolosclerosis of skeleton muscle is above all characterized by an increase of heterogeneous proteins (blood plasma proteins, actomyosin) in the arteriolar wall, leading to a diminution of the submicroscopical volume of gaps and to an elevation of the dry mass concentration.

Arteries↗
PubMed2003

Interaction of the mitotic inhibitor monastrol with human kinesin Eg5.

The microtubule-dependent kinesin-like protein Eg5 from Homo sapiens is involved in the assembly of the mitotic spindle. It shows a three-domain structure with an N-terminal motor domain, a central coiled coil, and a C-terminal tail domain. In vivo HsEg5 is reversibly inhibited by monastrol, a small cell-permeable molecule that causes cells to be arrested in mitosis. Both monomeric and dimeric Eg5 constructs have been examined in order to define the minimal monastrol binding domain on HsEg5. NMR relaxation experiments show that monastrol interacts with all of the Eg5 constructs used in this study. Enzymatic techniques indicate that monastrol partially inhibits Eg5 ATPase activity by binding directly to the motor domain. The binding is noncompetitive with respect to microtubules, indicating that monastrol does not interfere with the formation of the motor-MT complex. The binding is not competitive with respect to ATP. Both enzymology and in vivo assays show that the S enantiomer of monastrol is more active than the R enantiomer and racemic monastrol. Stopped-flow fluorometry indicates that monastrol inhibits ADP release by forming an Eg5-ADP-monastrol ternary complex. Monastrol reversibly inhibits the motility of human Eg5. Monastrol has no inhibitory effect on the following members of the kinesin superfamily: MC5 (Drosophila melanogaster Ncd), HK379 (H. sapiens conventional kinesin), DKH392 (D. melanogaster conventional kinesin), BimC1-428 (Aspergillus nidulans BimC), Klp15 (Caenorhabditis elegans C-terminal motor), or Nkin460GST (Neurospora crassa conventional kinesin).

Adenosine Diphosphate↗
PubMed1997

Identification of a mid-anaphase checkpoint in budding yeast.

Activation of a facultative, dicentric chromosome provides a unique opportunity to introduce a double strand DNA break into a chromosome at mitosis. Time lapse video enhanced-differential interference contrast analysis of the cellular response upon dicentric activation reveals that the majority of cells initiates anaphase B, characterized by pole-pole separation, and pauses in mid-anaphase for 30-120 min with spindles spanning the neck of the bud before completing spindle elongation and cytokinesis. The length of the spindle at the delay point (3-4 microm) is not dependent on the physical distance between the two centromeres, indicating that the arrest represents surveillance of a dicentric induced aberration. No mid-anaphase delay is observed in the absence of the RAD9 checkpoint gene, which prevents cell cycle progression in the presence of damaged DNA. These observations reveal RAD9-dependent events well past the G2/M boundary and have considerable implications in understanding how chromosome integrity and the position and state of the mitotic spindle are monitored before cytokinesis.

Anaphase↗
PubMed1997

Reconstruction of optical pathlength distributions from images obtained by a wide-field differential interference contrast microscope.

An image processing algorithm is presented to reconstruct optical pathlength distributions from images of nonabsorbing weak phase objects, obtained by a differential interference contrast (DIC) microscope, equipped with a charge-coupled device camera. The method is demonstrated on DIC images of transparent latex spheres and unstained bovine spermatozoa. The images were obtained with a wide-field DIC microscope, using monochromatic light. After image acquisition, the measured intensities were converted to pathlength differences. Filtering in the Fourier domain was applied to correct for the typical shadow-cast effect of DIC images. The filter was constructed using the lateral shift introduced in the microscope, and parameters describing the spectral distribution of the signal-to-noise ratio. By varying these parameters and looking at the resulting images, an appropriate setting for the filter parameters was found. In the reconstructed image each grey value represents the optical pathlength at that particular location, enabling quantitative analysis of object parameters using standard image processing techniques. The advantage of using interferometric techniques is that measurements can be done on transparent objects, without staining, enabling observations on living cells. Quantitative use of images obtained by a wide-field DIC microscope becomes possible with this technique, using relatively simple means.

Animals↗
PubMed1996

Quantitative analysis of cadherin-catenin-actin reorganization during development of cell-cell adhesion.

Epithelial cell-cell adhesion requires interactions between opposing extracellular domains of E-cadherin, and among the cytoplasmic domain of E-cadherin, catenins, and actin cytoskeleton. Little is known about how the cadherin-catenin-actin complex is assembled upon cell-cell contact, or how these complexes initiate and strengthen adhesion. We have used time-lapse differential interference contrast (DIC) imaging to observe the development of cell-cell contacts, and quantitative retrospective immunocytochemistry to measure recruitment of proteins to those contacts. We show that E-cadherin, alpha-catenin, and beta-catenin, but not plakoglobin, coassemble into Triton X-100 insoluble (TX-insoluble) structures at cell-cell contacts with kinetics similar to those for strengthening of E-cadherin-mediated cell adhesion (Angres, B., A. Barth, and W.J. Nelson. 1996. J. Cell Biol. 134:549-557). TX-insoluble E-cadherin, alpha-catenin, and beta-catenin colocalize along cell-cell contacts in spatially discrete micro-domains which we designate "puncta," and the relative amounts of each protein in each punctum increase proportionally. As the length of the contact increases, the number of puncta increases proportionally along the contact and each punctum is associated with a bundle of actin filaments. These results indicate that localized clustering of E-cadherin/catenin complexes into puncta and their association with actin is involved in initiating cell contacts. Subsequently, the spatial ordering of additional puncta along the contact may be involved in zippering membranes together, resulting in rapid strengthening of adhesion.

Actins↗
PubMed2004

Quality assurance considerations for detection of waterborne zoonotic parasites using Cryptosporidium oocyst detection as the main example.

A laboratory quality assurance (QA) program can minimize errors and provide confidence in the validity of laboratory test results. The structure of a QA program varies somewhat among laboratories but usually requires addressing a QA manual, QA goals, quality of resources, standard operating procedures, internal quality control, and external QA procedures. This paper reviews these general components and discusses some of the more particular QA considerations specific to filtration, immunomagnetic separation (IMS), immunofluorescence microscopy (FA), vital dye staining, differential interference contrast (DIC) microscopy, and molecular methods, which are involved in the detection and enumeration of Cryptosporidium oocysts.

Animals↗
PubMed2003

Near-field scanning optical microscopy imaging of multimode interference.

By use of a near-field scanning optical microscope (NSOM) in collection mode, the intensity distribution along a 2 x 2 multimode interference coupler was directly imaged as a function of wavelength. Although calculations can predict the general trend of wavelength dependence and the approximate positions of multiple images in the coupler, the accuracy is poor because of uncertainties in the waveguide width. We show that direct imaging using a NSOM bypasses calculational uncertainties and proves to be a powerful technique for studying these waveguide devices.

Journal Article↗
PubMed1976

Evaluation of bovine spermatozoal morphologic features after staining or fixation.

Two experiments were conducted to evaluate effects of 3 stains and 2 fixatives on morphologic features of bovine spermatozoa. In experiment 1, the morphologic features of acrosomes of raw and incubated, extended spermatozoa were evaluated after staining with Hancock's Blom's or Wells-Awa's stains or after fixation with buffered glutaraldehyde. Evaluations were done of stained smears by bright field microscopy and of fixed, unstained preparations, by differential interference contrast microscopy, using wet mounts. Raw semen samples from 1st ejaculates of 80 bulls were evaluated. The percentage of spermatozoa with intact acrosomes averaged 83.5% in unstained preparations fixed in glutaraldehyde, compared with averages of 68.1, 74.5, and 67.4% for smears stained with Hancock's, Blom's, or Wells-Awa's procedures (P less than 0.01). From these results, it appeared that procedures for preparing stained smears were detrimental to acrosomes. Although counts for other acrosomal abnormalities differed (P less than 0.01) in each treatment, patterns were inconsistent. With incubated, extended spermatozoa from 57 bulls, glutaraldehyde-fixed, unstained samples had more (55%) intact acrosomes (P less than 0.01) than did samples stained with Hancock's or Blom's procedures (24.0 and 34.7%, respectively, but the former were not significantly different from Wells-Awa-stained smears (49.3% intact acrosomes). In experiment 2, several morphologic characteristics of spermatozoa from 15 1st ejaculates of 7 bulls were evaluated after staining with Hancock's or Blom's stains or after fixation in buffered glutaraldehyde or buffered formal saline fixatives. Higher counts (P less than 0.01) of head abnormalities were found in wet, unstained fixed preparations (4.83, 4.47, 7.87, and 7.93% respectively, for Hancock's, Blom's, glutaraldehyde, and formol saline methods). There were more (P less than 0.05) separated heads on stained, dry smears (1.43, 1.23, 0.47, and 0.47%, respectively, for Hancock's, Blom's, glutaraldehyde, and formol saline procedures). Fixation with buffered glutaraldehyde resulted in higher counts (P less than 0.01) of proximal protoplasmic droplets (2.47, 1.03, 0.67, and 1.43%, respectively, for glutaraldehyde, Hancock's, Blom's, and formol saline procedures). Although not significant, the same trend was observed for distal protoplasmic droplets...

Acrosome↗