scholarly journals Epitope Mapping with Membrane-Bound Synthetic Overlapping Peptides

2014 ◽  
pp. 421-426
Author(s):  
Terumi Midoro-Horiuti ◽  
Randall M. Goldblum
Keyword(s):  
Epitope Mapping ◽  
Membrane Bound

scholarly journals A combined ELISA-immunoelectron microscopic approach for topological mapping of membrane protein epitopes: application to the nicotinic acetylcholine receptor.

1994 ◽  
Vol 42 (3) ◽  
pp. 315-327 ◽  
Author(s):  
M Schmutz ◽  
D Kling ◽  
S Tzartos ◽  
A Brisson
Keyword(s):  
Membrane Proteins ◽  
Acetylcholine Receptor ◽  
Epitope Mapping ◽  
Lipid Membrane ◽  
Torpedo Marmorata ◽  
Microscopic Approach ◽  
Electron Microscopic ◽  
Experimental Conditions ◽  
General Application ◽  
Membrane Bound

Identification of epitope localization on either side of the lipid membrane by immunoelectron microscopy constitutes an intrinsic powerful method of structure determination for membrane proteins. We have developed a method allowing measurement and observation, under almost identical experimental conditions, of the binding of monoclonal antibodies (MAb) to membrane-bound acetylcholine receptor from Torpedo marmorata electric tissue. This method, based on ELISA and electron microscopy of negatively stained specimens, was developed with MAb of known epitope specificity. With native membrane fragments, we found that MAb bound to extracellular epitopes in a stoichiometric manner, whereas almost no binding was detected for intracellular epitopes. The treatment based on tissue homogenization in the presence of Zn2+ ions and sucrose resulted in the formation of large, stable openings, rendering accessible about 25% of intracellular epitopes. Electron microscopic observations showed a clear distinction between antibody binding to either intracellular or extracellular epitopes, both with native and Zn(2+)-treated membranes. In addition, the binding of one antibody directed against an extracellular epitope was strikingly dependent on the packing density of acetylcholine receptor molecules, thus enabling us to further distinguish between two levels of accessibility for extracellular epitopes. The method presented here is of general application for studies of epitope mapping in membrane proteins.

Two Distinct Types of Microfilaments in the Cytoplasm of Human Adenohypophysial Cells

1973 ◽  
Vol 31 ◽  
pp. 668-669
Author(s):  
E. Horvath ◽  
K. Kovacs ◽  
I. E. Stratmann ◽  
C. Ezrin
Keyword(s):  
Pituitary Tumors ◽  
Average Diameter ◽  
Anterior Lobe ◽  
Uranyl Acetate ◽  
Type I ◽  
Breast Cancer Patients ◽  
Human Pituitary ◽  
Severe Diabetes ◽  
Pituitary Glands ◽  
Membrane Bound

Surgically removed human pituitary glands as well as pituitary tumors fixed in glutaraldehyde, postfixed in osmium tetroxide, embedded in epon resin, stained with uranyl acetate and lead citrate have been investigated by electron microscopy in order to correlate ultrastructure with functional activity. In the course of this study two distinct types of microfilaments have been identified in the cytoplasm of adenohypophysiocytes.Type I microfilaments (Fig. 1) were found in the cytoplasm of anterior lobe cells of five female subjects with disseminated mammary cancer and two patients with severe diabetes mellitus. The breast cancer patients were treated pre-operatively for various periods of time with different doses of oxysteroids. The microfilaments had an average diameter of JO A, formed parallel bundles, were scattered irregularly in the cytoplasm and were frequently located in the perikaryon. They were not membrane-bound and failed to show any periodicity.

Intranuclear Crystals in Regenerating Canine Kidneys

1973 ◽  
Vol 31 ◽  
pp. 412-413
Author(s):  
D.G. Osborne ◽  
L.J. McCormack ◽  
M.O. Magnusson ◽  
W.S. Kiser
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Tubular Epithelial Cell ◽  
Tubular Epithelial Cells ◽  
Cell Nuclei ◽  
Crystalline Structures ◽  
Small Crystals ◽  
Membrane Bound

During a project in which regenerative changes were studied in autotransplanted canine kidneys, intranuclear crystals were seen in a small number of tubular epithelial cells. These crystalline structures were seen in the control specimens and also in regenerating specimens; the main differences being in size and number of them. The control specimens showed a few tubular epithelial cell nuclei almost completely occupied by large crystals that were not membrane bound. Subsequent follow-up biopsies of the same kidneys contained similar intranuclear crystals but of a much smaller size. Some of these nuclei contained several small crystals. The small crystals occurred at one week following transplantation and were seen even four weeks following transplantation. As time passed, the small crystals appeared to fuse to form larger crystals.

Membrane-bound vesicles associated with the microvilli in the midgut of the freshwater microcrustacean, Daphnia magna

1983 ◽  
Vol 41 ◽  
pp. 480-481
Author(s):  
Patricia L. Jansma
Keyword(s):  
Daphnia Magna ◽  
Intestinal Epithelial Cells ◽  
Uranyl Acetate ◽  
Intestinal Epithelial ◽  
Chlamydomonas Reinhardii ◽  
Thin Sections ◽  
Saturated Water ◽  
Cacodylate Buffer ◽  
Membrane Bound ◽  
Ethanol Series

The presence of the membrane bound vesicles or blebs on the intestinal epithelial cells has been demonstrated in a variety of vertebrates such as chicks, piglets, hamsters, and humans. The only invertebrates shown to have these microvillar blebs are two species of f1ies. While investigating the digestive processes of the freshwater microcrustacean, Daphnia magna, the presence of these microvillar blebs was noticed.Daphnia magna fed in a suspension of axenically grown green alga, Chlamydomonas reinhardii for one hour were narcotized with CO2 saturated water. The intestinal tracts were excised in 2% glutaraldehyde in 0.2 M cacodyl ate buffer and then placed in fresh 2% glutaraldehyde for one hour. After rinsing in 0.1 M cacodylate buffer, the sample was postfixed in 2% OsO4, dehydrated with a graded ethanol series, infiltrated and embedded with Epon-Araldite. Thin sections were stained with uranyl acetate and Reynolds lead citrate before viewing with the Philips EM 200.

Analysis of the Anterior Secretory Cells of Onchidohs Muricata (Nudibranchia)

1981 ◽  
Vol 39 ◽  
pp. 614-615
Author(s):  
Roy Skidmore
Keyword(s):  
Endoplasmic Reticulum ◽  
Secretory Granules ◽  
Golgi Body ◽  
The Body ◽  
Secretory Cells ◽  
Secretory Vesicles ◽  
High Magnification ◽  
Large Numbers ◽  
Membrane Bound ◽  
Sole Of The Foot

The long-necked secretory cells in Onchidoris muricata are distributed in the anterior sole of the foot. These cells are interspersed among ciliated columnar and conical cells as well as short-necked secretory gland cells. The long-necked cells contribute a significant amount of mucoid materials to the slime on which the nudibranch travels. The body of these cells is found in the subepidermal tissues. A long process extends across the basal lamina and in between cells of the epidermis to the surface of the foot. The secretory granules travel along the process and their contents are expelled by exocytosis at the foot surface.The contents of the cell body include the nucleus, some endoplasmic reticulum, and an extensive Golgi body with large numbers of secretory vesicles (Fig. 1). The secretory vesicles are membrane bound and contain a fibrillar matrix. At high magnification the similarity of the contents in the Golgi saccules and the secretory vesicles becomes apparent (Fig. 2).

An Electron Microscopic Study of the Acute Effects of Hypothalamic Releasing Factors on the Anterior Pituitary Gland

1968 ◽  
Vol 26 ◽  
pp. 232-233
Author(s):  
P.W. Coates ◽  
E.A. Ashby ◽  
L. Krulich ◽  
A. Dhariwal ◽  
S. McCann
Keyword(s):  
Anterior Pituitary ◽  
Microscopic Investigation ◽  
Uranyl Acetate ◽  
Electron Microscopic Investigation ◽  
Electron Microscopic ◽  
Tissue Samples ◽  
Thin Sections ◽  
Dense Membrane ◽  
Membrane Bound ◽  
Releasing Factors

The morphologic effects on somatotrophs of crude sheep hypothalamic extract prepared from stalk-median eminence were studied by electron microscopy in conjunction with concurrently run bioassays performed on the same tissue samples taken from young adult male Sherman rats.Groups were divided into uninjected controls and injected experimentals sacrificed at 5', 15', and 30' after injection. Half of each anterior pituitary was prepared for electron microscopic investigation, the other half for bioassay. Fixation using collidine buffered osmium tetroxide was followed by dehydration and embedment in Maraglas. Uranyl acetate and lead citrate were used as stains. Thin sections were examined in a Philips EM 200.Somatotrophs from uninjected controls appeared as described in the literature (Fig. 1). In addition to other components, these cells contained moderate numbers of spherical, electron-dense, membrane-bound granules approximately 350 millicrons in diameter.

A High Resolution Cytochemical Study of Nuclear ATPase in Human Spermatozoa

1975 ◽  
Vol 33 ◽  
pp. 594-595
Author(s):  
A. Sosa ◽  
L. Calzada
Keyword(s):  
High Resolution ◽  
Atpase Activity ◽  
Nuclear Membrane ◽  
Human Sperm ◽  
Sperm Nucleus ◽  
Human Spermatozoa ◽  
Cytochemical Study ◽  
Membrane Bound ◽  
Sperm Nuclei ◽  
Interchromatin Space

The dependence of nuclear metabolism on the function of the nuclear membrane is not well understood. Whether or not the function of the nuclear membrane is partial or totally responsible of the repressed template activity of human sperm nucleus has not at present been elucidated. One of the membrane-bound enzymatic activities which is concerned with the mechanisms whereby substances are thought to cross cell membranes is adenosintriphosphatase (ATPase). This prompted its characterization and distribution by high resolution photogrammetry on isolated human sperm nuclei. Isolated human spermatozoa nuclei were obtained as previously described. ATPase activity was demonstrated by the method of Wachstein and Meisel modified by Marchesi and Palade. ATPase activity was identified as dense and irregularly distributed granules confined to the internal leaflet of the nuclear membrane. Within the nucleus the appearance of the reaction product occurs as homogenous and dense precipitates in the interchromatin space.

Spindle shaped bodies in foveolar cones of the baboon retina

1989 ◽  
Vol 47 ◽  
pp. 960-961
Author(s):  
W. Krebs ◽  
I. Krebs
Keyword(s):  
Cross Sections ◽  
Inclusion Bodies ◽  
Photoreceptor Cells ◽  
Papio Anubis ◽  
Retinal Photoreceptor ◽  
Age Related ◽  
Cone Cells ◽  
Membrane Bound ◽  
Oriented Parallel ◽  
Shaped Inclusion

Various inclusion bodies occur in vertebrate retinal photoreceptor cells. Most of them are membrane bound and associated with phagocytosis or they are age related residual bodies. We found an additional inclusion body in foveal cone cells of the baboon (Papio anubis) retina.The eyes of a 15 year old baboon were fixed by immersion in cacodylate buffered glutaraldehyde (2%)/formaldehyde (2%) as described in detail elsewhere . Pieces of retina from various locations, including the fovea, were embedded in epoxy resin such that radial or tangential sections could be cut.Spindle shaped inclusion bodies were found in the cytoplasm of only foveal cones. They were abundant in the inner segments, close to the external limiting membrane (Fig. 1). But they also occurred in the outer fibers, the perikarya, and the inner fibers (Henle’s fibers) of the cone cells. The bodies were between 0.5 and 2 μm long. Their central diameter was 0.2 to 0. 3 μm. They always were oriented parallel to the long axis of the cone cells. In longitudinal sections (Figs. 2,3) they seemed to have a fibrous skeleton that, in cross sections, turned out to consist of plate-like (Fig.4) and tubular profiles (Fig. 5).

The ultrastructure of the double membrane-bound bodies and endoplasmic reticulum in serial sections of wheat spot mosaic-affected wheat plants

1990 ◽  
Vol 48 (3) ◽  
pp. 694-695
Author(s):  
M. H. Chen ◽  
C. Hiruki
Keyword(s):  
Endoplasmic Reticulum ◽  
High Resolution ◽  
Membrane Structure ◽  
Mosaic Disease ◽  
Serial Sections ◽  
Thin Sections ◽  
Double Membrane ◽  
Southern Alberta ◽  
Membrane Bound ◽  
Scanning Em

Wheat spot mosaic disease was first discovered in southern Alberta, Canada, in 1956. A hitherto unidentified disease-causing agent, transmitted by the eriophyid mite, caused chlorosis, stunting and finally severe necrosis resulting in the death of the affected plants. Double membrane-bound bodies (DMBB), 0.1-0.2 μm in diameter were found to be associated with the disease.Young tissues of leaf and root from 4-wk-old infected wheat plants were fixed, dehydrated, and embedded in Spurr’s resin. Serial sections were collected on slot copper grids and stained. The thin sections were then examined with a Hitachi H-7000 TEM at 75 kV. The membrane structure of the DMBBs was studied by numbering them individually and tracing along the sections to see any physical connection with endoplasmic reticulum (ER) membranes. For high resolution scanning EM, a modification of Tanaka’s method was used. The specimens were examined with a Hitachi Model S-570 SEM in its high resolution mode at 20 kV.

Portein-gold labelling of ultrathin sections of wheat spot mosaic-affected wheat plants

1990 ◽  
Vol 48 (3) ◽  
pp. 680-681
Author(s):  
K. S. Zaychuk ◽  
M. H. Chen ◽  
C. Hiruki
Keyword(s):  
Osmium Tetroxide ◽  
Rnase A ◽  
Leaf Ultrastructure ◽  
Double Membrane ◽  
Young Root ◽  
Leaf Tissues ◽  
Membrane Bound ◽  
Ultrathin Sections ◽  
Gold Labelling ◽  
Electron Dense Core

Wheat spot mosaic (WSpM), which frequently occurs with wheat streak mosaic virus was first reported in 1956 from Alberta. Singly isolated, WSpM causes chlorotic spots, chlorosis, stunting, and sometimes death of the wheat plants. The vector responsible for transmission is the eriophyid mite, Eriophyes tulipae Kiefer. The examination of leaf ultrastructure by electron microscopy has revealed double membrane bound bodies (DMBB’s) 0.1-0.2 μm in diameter. Dispersed fibrils within these bodies suggested the presence of nucleic acid. However, neither ribosomes characteristic of bacteria, mycoplasma and the psittacosis group of organisms nor an electron dense core characteristic of many viruses was commonly evident.In an attempt to determine if the DMBB’s contain nucleic acids, RNase A, DNase I, and lactoferrin protein were conjugated with 10 nm colloidal gold as previously described. Young root and leaf tissues from WSpM-affected wheat plants were fixed in glutaraldehyde, postfixed in osmium tetroxide,and embedded in Spurr’s resin.

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