scholarly journals Purification of an 80,000-dalton protein that is a component of the isolated microvillus cytoskeleton, and its localization in nonmuscle cells.

1983 ◽  
Vol 97 (2) ◽  
pp. 425-432 ◽  
Author(s):  
A Bretscher
Keyword(s):  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Cultured Cells ◽  
Major Protein ◽  
Intestinal Epithelial ◽  
Indirect Immunofluorescence Microscopy ◽  
Core Proteins ◽  
Nonmuscle Cells ◽  
Protein Components

The microvillus cytoskeleton, isolated from chicken intestinal epithelial cell brush borders, is known to contain five major protein components, the 110,000-dalton polypeptide, villin (95,000 daltons), fimbrin (68,000 daltons), actin (43,000 daltons), and calmodulin (17,000 daltons). In this paper we describe our first step in studying the minor components of the isolated core. We have so far identified and purified an 80,000-dalton polypeptide that was present in the isolated structure in approximately 0.7% the molar abundance of actin. Antibodies to the 80,000-dalton component did not react with other microvillus core proteins, and, when used in indirect immunofluorescence microscopy, they stained the microvilli of intestinal epithelial cells fixed in situ. The 80,000-dalton component therefore appears to be a newly-identified, authentic component of intestinal microvilli in vivo and of isolated microvillus cores. Immunological studies demonstrate that the 80,000-dalton component is widely distributed in nonmuscle cells. Indirect immunofluorescence microscopy reveals that it is particularly enriched in surface structures, such as blebs, microvilli, and retraction fibers of cultured cells.

scholarly journals Fimbrin, a new microfilament-associated protein present in microvilli and other cell surface structures.

1980 ◽  
Vol 86 (1) ◽  
pp. 335-340 ◽  
Author(s):  
A Bretscher ◽  
K Weber
Keyword(s):  
Cell Surface ◽  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Cultured Cells ◽  
Intestinal Epithelial Cells ◽  
Surface Structures ◽  
Intestinal Epithelial ◽  
Indirect Immunofluorescence Microscopy ◽  
Cell Surface Structures ◽  
Microfilament Bundles

A 68,000 mol wt polypeptide has been identified as one of the few major proteins in the microfilament bundles of the microvilli present on intestinal epithelial cells. Antibodies against the purified protein have been used in indirect immunofluorescence microscopy on several cultured cells. The protein have been used in indirect immunofluorescence microscopy on several cultured cells. The protein is found particularly prominent in membrane ruffles, microspikes, and microvilli.

scholarly journals Localization of actin and microfilament-associated proteins in the microvilli and terminal web of the intestinal brush border by immunofluorescence microscopy.

1978 ◽  
Vol 79 (3) ◽  
pp. 839-845 ◽  
Author(s):  
A Bretscher ◽  
K Weber
Keyword(s):  
Epithelial Cells ◽  
Indirect Immunofluorescence ◽  
Brush Border ◽  
Immunofluorescence Microscopy ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Indirect Immunofluorescence Microscopy ◽  
Terminal Web ◽  
Intestinal Brush Border ◽  
Associated Proteins

Indirect immunofluorescence microscopy was used to localize microfilament-associated proteins in the brush border of mouse intestinal epithelial cells. As expected, antibodies to actin decorated the microfilaments of the microvilli, giving rise to a very intense fluorescence. By contrast, antibodies to myosin, tropomyosin, filamin, and alpha-actinin did not decorate the microvilli. All these antibodies, however, decorated the terminal web region of the brush border. Myosin, tropomyosin, and alpha-actinin, although present throughout the terminal web, were found to be preferentially located around the periphery of the organelle. Therefore, two classes of microfilamentous structures can be documented in the brush border. First, the highly ordered microfilaments which make up the cores of the microvilli apparently lack the associated proteins. Second, seemingly less-ordered microfilaments are found in the terminal web, in which region the myosin, tropomyosin, filamin and alpha-actinin are located.

Syncytioskeletons in choriocarcinoma in culture

1984 ◽  
Vol 66 (1) ◽  
pp. 1-20
Author(s):  
C.D. Ockleford ◽  
L. Dearden ◽  
R.A. Badley
Keyword(s):  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Cultured Cells ◽  
Cortical Layers ◽  
Dense Population ◽  
Bewo Cells ◽  
Indirect Immunofluorescence Microscopy ◽  
Microtubule Organizing Centres ◽  
Transmission Electron ◽  
Cytoskeletal Elements

Indirect immunofluorescence microscopy using anti-actin serum has been used to investigate the distribution of actin-containing polymers in BeWo cells. This cell line, derived from a human choriocarcinoma, contains tissue that, like its tissue of origin, is partly syncytial. The syncytial nature has been inferred from study of Nomarski optical sections and from transmission electron microscopy. The multinucleated plaques of tissue possess a syncytioskeleton with a number of actin-containing features characteristic of cultured cells. These include stress fibres, cortical layers and ruffled membranes. Other actin-containing structures are more typical of the related non-pathological syncytiotrophoblast. These include a dense population of microvilli. The overall organization of the actin syncytioskeletons bears no obvious relationship to the number or position of nuclei in the syncytium. Indirect immunofluorescence microscopy has also been employed to localize the protein tubulin in BeWo cells. The microtubules do not appear to be spatially organized by a particular nucleus. Rather, there are numerous microtubule-organizing centres (MTOCs) that exist in the cytoplasm and do not have the expected numerical and positional relationship to nuclei. From these data it appears that polymeric cytoskeletal elements in these syncytia are organized in a manner not immediately subordinate to syncytial nuclei.

scholarly journals Intermediate-sized filaments of human endothelial cells.

1979 ◽  
Vol 81 (3) ◽  
pp. 570-580 ◽  
Author(s):  
W W Franke ◽  
E Schmid ◽  
M Osborn ◽  
K Weber
Keyword(s):  
Electron Microscopy ◽  
Endothelial Cells ◽  
Smooth Muscle ◽  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Major Protein ◽  
Human Endothelial Cells ◽  
Specific Antibodies ◽  
Indirect Immunofluorescence Microscopy ◽  
Polypeptide Band

Human endothelial cells prepared from unbilical cords are characterized in parallel by electron microscopy and indirect immunofluorescence microscopy using specific antibodies against different classes of intermediate-sized filaments. The strongly developed, loose bundles of intermediate-sized filaments typically found in these cells are not decorated by antibodies against prekeratin or antibodies against smooth muscle desmin. They are, however, strongly decorated by antibodies directed against murine "vimentin," i.e., the 57,000 mol wt polypeptide which is the major protein of the intermediate-sized filaments predominant in various cells of mesenchymal origin. Cytoskeletal preparations greatly enriched in intermediate-sized filaments show the enrichment of a polypeptide band comigrating with murine vimentin. This shows that the intermediate-sized filaments that are abundant in human endothelial cells are predominantly of the vimentin type and can be demonstrated by their cross-reaction with the vimentin of rodents. These data also strengthen the evidence for several subclasses of intermediate-sized filaments, which can be distinguished by immunological procedures.

scholarly journals Binding of deoxyribonuclease I to actin: a new way to visualize microfilament bundles in nonmuscle cells.

1978 ◽  
Vol 26 (9) ◽  
pp. 745-749 ◽  
Author(s):  
E Wang ◽  
A R Goldberg
Keyword(s):  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Staining Pattern ◽  
Fluorescent Microscopy ◽  
Dnase I ◽  
Deoxyribonuclease I ◽  
Indirect Immunofluorescence Microscopy ◽  
Microfilament Bundles ◽  
Nonmuscle Cells ◽  
In Cells

Intracellular actin-containing fibers can be visualized by indirect immunofluorescence microscopy when they are stained with antibody directed against DNase I. The location of actin-containing fibers in cells appears to be similar to the staining pattern of antibody to actin. Actin fibers were also visualized by direct fluorescent microscopy with rhodamine-conjugated DNase I.

scholarly journals Localization of protein disulfide isomerase to the external surface of the platelet plasma membrane

Blood ◽  
1995 ◽  
Vol 86 (6) ◽  
pp. 2168-2173 ◽  
Author(s):  
DW Essex ◽  
K Chen ◽  
M Swiatkowska
Keyword(s):  
Indirect Immunofluorescence ◽  
Blood Cells ◽  
Disulfide Bonds ◽  
Protein Disulfide Isomerase ◽  
Immunofluorescence Microscopy ◽  
External Surface ◽  
Platelet Surface ◽  
Disulfide Isomerase ◽  
Indirect Immunofluorescence Microscopy ◽  
Protein Disulfide

Protein disulfide isomerase (PDI) is an enzyme that catalyzes the formation as well as the isomerization of disulfide bonds. In this study, antibodies against PDI were used to show PDI antigen on the platelet surface by indirect immunofluorescence microscopy and by flow cytometry. The platelets were not activated, as evidenced by the absence of staining by an antibody against P-selectin. Permeabilized platelets showed little cytosolic PDI by indirect immunofluorescence microscopy, suggesting that the majority of platelet PDI is localized to the platelet surface. PDI activity against “scrambled” RNase was shown with intact platelets. The activity was inhibited by inhibitors of PDI and by an antibody against PDI. Other blood cells showed little PDI. Platelet surface PDI may play a role in the various physiological and pathophysiologic processes in which platelets are involved.

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