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  • Life and Medical Sciences  (1,377)
  • 1
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 26 (1990), S. 175-183 
    ISSN: 1040-452X
    Keywords: Sperm antigen ; Testis ; Blood-testis ; barrier ; Antisperm antibodies ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The fertiization antigen (FA-1) isolated from murine testes demonstrated its dimeric form of 49,000 ± 2,000 molecular weight (M.W.) or a monomer of 23,000 M.W. on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The FA-1 was immunogenic in all three female rabbits tested and raised a high-titer antisera [enzyme-linked immunosorbent assay (ELISA) titers; 1:1,024 to 1:4,096]. The rabbit anti-FA-1 antisera predominantly recognized the dimeric form of 49,000 ± 2,000 M.W. on the Western blot of lithium diiodosalicylate (LIS)-solubilized murine testes. None of the antisera reacted with any somatic tissue, indicating germ-cell specificity of FA-1. To determine the cellular localization of the immunoreactive FA-1, a novel ultrasensitive immunogold-silver staining (IGSS) procedure was developed. The anti-FA-1-lgG showed intense staining in the luminal region of the seminiferous tubules containing spermatids and spermatozoa. No reaction was observed in the peripheral area of the tubules containing Sertoli cells, spermatogonia, leptotene, and zygotene spermatocytes. The biodistribution studies of 125I-labeled anti-FA-1 lgG in mice revealed that the antibodies do not bind to somatic tissues such as blood cell, liver, heart, kidney, muscle, and gastrointestinal tissue and do not transudate into testes nad seminal vesicle. However, the antibodies preferentially transudate into epididymis (especially corpus or cauda regions) and vas deferens to bind to sperm cells. In conclusion, our data indicate that FA-1 can induce an immune response that is germ cell-specific, directed against later stages of spermatogenesis. The antibodies to FA-1 interact with sperm after penetration through epididymis (especially corpus and cauda regions) and vas deferens rather than through testes and seminal vesicles.
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  • 2
    Electronic Resource
    Electronic Resource
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 37 (1988), S. 193-202 
    ISSN: 0730-2312
    Keywords: colorectal carcinoma ; Carcinoembryonic antigen ; UEA ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Recent interest has focused on fucosylated epitopes expressed on human neoplasms. The plant lectin Ulex europus agglutinin, Type I (UEA) binds fucosylated oligosac-charides, while UEA-reactive substances have a tissue distribution similar to carci-noembryonic antigen (CEA). We sought to determine if UEA reacted with CEA in extracts of fresh primary and metastatic colorectal carcinomas and paired normal tissues. The extracts were electrophoretically transferred to nitrocellulose membranes after the proteins were separated by SDS-PAGE in 10% polyacrylamide gels. The transfer membranes were then stained with peroxidase-conjugated UEA (UEA-P) or antibody to CEA (CEA-P). UEA-P reacted with a 170-190-kDa band in extracts of 22 of 30 primary tumors, 10 of 12 metastases, but only 1 of 5 villous adenomas. UEA-P generally did not react with normal colon or liver extracts. UEA-P also did not bind to 170-190-kDa molecules in Western transfers of a breast carcinoma metastatic to bowel and a focal nodular hyperplasia of liver. CEA-P displayed similar reactivity and detected CEA in a tumor extract negative for UEA. Fucose blocked binding of UEA-P to Western transfers of tumor extracts. CEA-P reacted with a 170-190-kDa substance in tumor extracts eluted with fucose from a column of immobilized UEA. Thus, UEA reacts with fucosylated oligosaccharides on most, but not all, species of CEA and may be a useful adjunct to anti-CEA immunohis-tochemistry.
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Journal of Electron Microscopy Technique 2 (1985), S. 139-146 
    ISSN: 0741-0581
    Keywords: Immunoelectron microscopy ; colloidal gold ; localization of proteins ; Sendai virus nucleocapsid ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Natural Sciences in General
    Notes: We have described an indirect immunoelectron microscope method to localize individual proteins on viral nucleocapsids using monoclonal antibodies and colloidal gold-conjugated second antibodies. The procedure provides good binding and retention of antibodies, good resolution(〈 24nm), and negligible nonspecific binding of antibodies to the background. In addition, the method is compatible with both negative and positive staining, the two staining procedures commonly used to derive ultrastructural information on viral chromosomes. We have illustrated the procedure by localizing the NP (nucleoprotein, ≍ 2,600 copies) and P (polymerase-associated protein, ≍ 300 copies) proteins on the nucleocapsid of Sendai virus, a paramyxovirus.
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 20 (1991), S. 30-37 
    ISSN: 0886-1544
    Keywords: chick brain ; α-isotypes ; tyrosine ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The tyrosinylation of chick brain α-tubulin and the effects of the tyrosinylation status on the assembly and dynamic instability of chick brain MAP2:tubulin microtubule protein have been examined. Each of the eight major α-isotypes can be tyrosinylated in vitro, irrespective of whether a C-terminal tyrosine is genetically encoded. The extent of tyrosinylation is however limited to ≏ 0.3 mol.mol-1. The tyrosinylation status (0 vs. 0.3 mol.mol-1) has no effect on either the assembly kinetics of chick brain microtubule protein or on the rate of length redistribution following assembly and shearing. It is therefore unlikely that the tyrosinylation status directly affects the intrinsic stability of assembled microtu-bules since the rate of length redistribution is both a sensitive assay and a function of the kinetic parameters governing dynamic instability.
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  • 5
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Hepatocytes maintained in collagen gels remain differentiated for prolonged periods of time compared to cells maintained on conventional cultures. Previous studies with other culture systems in which chemical supplements or substratum modifications enhanced hepatocyte differentiation showed that in all of these systems hepatocytes do not respond to mitogens. In this study it is shown that hepatocytes maintained between two layers of collagen gels respond to mitogens HGF (also known as scatter factor (HGF/SF)) and epidermal growth factor (EGF). Cell density did not affect the responsiveness to mitogens as in conventional cultures. In addition both mitogens (HGF more pronounced) induce characteristic morphogenic changes in which hepatocytes form processes and join in formation of cords. Hepatocytes respond to mitogens for up to 6 days in culture at which point they become refractory to further mitogenic stimulation. This occurs despite electron microscopic evidence that these cells are fully viable when they become refractory to mitogenesis. The refractory state is not modified by substitution of one growth factor for the other or by addition of growth factors at different times. Hepatocytes in the refractory state become again responsive to mitogens when the collagen gels are dispersed by collagenase and the cells are replated on conventional substrates. © 1993 Wiley-Liss, Inc.
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  • 6
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Our laboratory has recently demonstrated that 1,25-dihydroxyvitamin D3(1,25(OH)2D3) rapidly stimulated membrane polyphosphoinositide breakdown and increased intracellular calcium, as well as activated protein kinase C (PKC) in vitamin D-sufficient rat colonocytes. These effects of 1,25(OH)2D3 were, however, lost in vitamin D-insufficient rats and restored by the in vivo repletion of 1,25(OH)2D3. In the present studies we have examined the ability of 1,25(OH)2D3 to stimulate the phosphorylation of colonic membrane proteins in intact D-sufficient cells. In addition, we investigated the effects of vitamin D status on the phosphorylation of these membrane proteins in broken cell preparations. These studies demonstrated that 1,25(OH)2D3 increased the phosphorylation of at least two colonic membrane proteins with apparent molecular weights of 42,000 (pp42) and 48,000 (pp48) in intact cells of vitamin D-sufficient rats. Moreover, in vitamin D-sufficient rats, treatment of colonocytes with 1,25(OH)2D3 or 12-Otertradecanoyl phorbol 13-acetate (TPA), a known activator of PKC, significantly increased the phosphorylation of pp42 and pp48 in broken cell preparations. The kinetics of these phosphorylations in response to 1,25(OH)2D3 were both rapid and transient. In addition, PKC19-36, a specific PKC inhibitor, decreased the phosphorylation of pp42 and pp48, whereas okadaic acid (OA), a type 1 and 2A protein phosphatase inhibitor, further augmented their phosphorylation in response to 1,25(OH)2D3. The isoelectric points of pp42 and pp48 were 5.79 and 5.97, respectively, and both were predominantly phosphorylated on threonine residues. In contrast to our findings in colonocytes from vitamin D-sufficient animals, basal phosphorylation of pp42 and pp48 were increased in membranes prepared from vitamin D-insufficient rats. Moreover, these phosphorylations failed to change in response to 1,25(OH)2D3-treatment of colonocytes from vitamin D-insufficient rats. The basal phosphorylation of each of these proteins was restored to control levels, as was their ability to respond to the direct addition of 1,25(OH)2D3 following the in vivo repletion of vitamin D-insufficient rats with this secosteroid. In summary, we have identified two acidic membrane proteins from rat colonocytes that are phosphorylated in both intact and broken cell preparations in response to 1,25(OH)2D3 treatment, an event modulated by vitamin D status and mediated, at least in part, by PKC. © 1995 Wiley-Liss, Inc.
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  • 7
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 157 (1993), S. 243-252 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The aim of this study was to determine the role of ECM components of bone in regulating the differentiation and function of cells of the osteoblast lineage. Rat UMR 201 cells, phenotypically preosteoblast, were plated onto plastic tissue culture dishes or dishes coated with gelled type I collagen or reconstituted basement membrane (matrigel). Acute cell attachment assays showed that cells adhered to substrates in the following order: collagen 〉 matrigel ≫ plastic. Proliferation rate up to 96 hr were similar on each substrate. However, if cells were treated with 10-6 M retinoic acid (RA), proliferation rates were reduced compared with control for cells grown on collagen and matrigel but not on plastic. Morphological changes were matrix-specific; in subconfluent cultures, long thin processes were seen with cells grown on collagen and a pattern of interconnecting cell processes formed when cells were plated on matrigel. Striking differences were observed in the constitutive or RA-induced gene expression of cells grown on the different substrates. When cells plated on collagen were treated with RA, induction of mRNA for alkaline phosphatase (ALP) as well as ALP enzyme activity were much less than with cells grown on plastic. In contrast, RA treatment induced osteopontin (OP) mRNA expression more strongly in cells plated on collagen compared with plastic within 24 hr and this was maintained for 72 hr. RA treatment produced a two fold increase of pro-α 1(I) collagen mRNA in cells grown on plastic and matrigel but not in cells grown on collagen. Growth on collagen produced changes in the way UMR 201 cells responded to RA from which they did not fully recover in subsequent 48-hr growth periods on plastic. These results indicate that ECM components regulate the function of and are capable of modulating RA-induced differentiation of preosteoblasts. © 1993 Wiley-Liss, Inc.
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  • 8
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 159 (1994), S. 229-237 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Programmed cell death is an active process wherein the cell initiates a sequence of events culminating in the fragmentation of its DNA, nuclear collapse, and disintegration of the cell into small, membrane-bound apoptotic bodies. Examination of the death program in various models has shown common themes, including a rise in cytoplasmic calcium, cytoskeletal changes, and redistribution of membrane lipids. The calcium-dependent neutral protease calpain has putative roles in cytoskeletal and membrane changes in other cellular processes; this fact led us to test the role of calpain in a well-known model of apoptotic cell death, that of thymocytes after treatment with dexamethasone. Assays for calcium-dependent proteolysis in thymocyte extracts reveal a rise in activity with a peak at about 1 hr of incubation with dexamethasone, falling to background at approximately 2 hr. Western blots indicate autolytic cleavage of the proenzyme precursor to the calpain I isozyme, providing additional evidence for calpain activation. We have also found that apoptosis in thymocytes, whether induced by dexamethasone or by low-level irradiation, is blocked by specific inhibitors of calpain. Apoptosis of metamyelocytes incubated with cycloheximide is also blocked by calpain inhibitors. These studies suggest a required role for calpain in both “induction” and “release” models of apoptotic cell death. © 1994 wiley-Liss, Inc.
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  • 9
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 146 (1991), S. 164-169 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The formation of blood vessels in vivo (angiogenesis) is an important process and is usually initiated in response to injury, tumor growth, or normal tissue development. We have studied the effect of human interferon (IFN) alpha (α) and gamma (γ) on the capillary-like network formation in an in vitro model of angiogenesis using human umbilical vein endothelial cells (HUVEC). When HUVEC cells are plated on Matrigel (reconstituted basement membrane matrix enriched in laminin), a network of capillary like structures (endotubes) rapidly forms. IFN-α enhanced the tube formation in a dose-dependent manner, whereas IFN-γ significantly inhibited the tube formation. In addition, both the enhancement and inhibition of angiogenesis by IFN-α and γ was found to be greater if the cells were pretreated with IFN for 12 hr before plating on the Matrigel. These results suggest that IFN may play an important role in several vascular processes including early stages of wound healing, recanalization of thrombi, tumor growth, metastasis, normal growth, and development.
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  • 10
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 147 (1991), S. 292-297 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A sialoglycopeptide (SGP), isolated and purified from bovine cerebral cortex cells, was studied in regard to early signal transduction events associated with the cell cycle. Previously shown to be a potent antagonist to a variety of mitogens, the SGP abrogated the ability of 12-O-tetradecanoylphorbol-13 acetate (TPA) to elicit an alkalinization of 3T3 cell cytosol, but only when added minutes prior to, or simultaneously with, the tumor promoter. 3T3 cell TPA-mediated Ca2+ mobilization was also inhibited by the SGP although the inhibitor itself did not bind Ca2+ in a cell-free assay. The results are discussed in light of the already known kinetics of interaction between the SGP, various mitogens, and the calcium ionophore A23187 with regard to the pivotal events leading to the decision of a cell to divide or not to divide.
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