Immunocytochemical demonstration of the binding and internalization of growth hormone, B-cell stimulating factor-1 and thy-1.2 in murine lymphocytes.

Wang, Jaang-Jiun; Liu, Hwan-Wun; Mao, Shou-Hsian; Yen, Ching‐Yu; Chang, Jeffrey P.

doi:10.1267/ahc.22.77

Acta Histochem. Cytochem

1989

DOI: 10.1267/ahc.22.77

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Immunocytochemical demonstration of the binding and internalization of growth hormone, B-cell stimulating factor-1 and thy-1.2 in murine lymphocytes.

Jaang-Jiun Wang

Hwan-Wun Liu

Shou-Hsian Mao

et al.

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1989

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Cited by 3 publications

References 28 publications

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Internalization of CD4 molecules in human T-cells demonstrated by immuno-electron microscopy

Wang

Lee³

et al. 1992

Histochemistry

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Internalization of CD4 molecules on human CD4-enriched T-cells was demonstrated by immunocytochemical electron microscopy. CD4+ T-cell subclones were obtained from normal human peripheral blood, followed by one-way MLC screening and co-culturing with IL-2. Fixed and non-fixed T-cell samples were indirectly immunolabeled with mouse anti-human CD4 monoclonal antibody and goat anti-mouse IgG conjugated with peroxidase. Unfixed T-cells were immunolabeled at 4 degrees C and then re-incubated for 5-45 min at 37 degrees C. The selected CD4+ T-cell subclones showed strong CD4 binding on the cell surface after IL-2 incubation. However, fresh T-cells, monocytes, bone marrow cells and CD8+ T-cells all stained negative for CD4. The distribution of CD4 molecules on the fixed cell surface showed a homogeneous pattern. Capping and internalization of CD4-antibody-peroxidase complexes from the cell surfaces were observed follow a pathway of receptor-mediated endocytosis in unfixed T cells. Endocytotic vesicles, vacuoles of diverse sizes and shapes near the cell membrane or deep in the cell center were found to contain CD4 molecules. Negatively stained Golgi saccules were observed up to 45 min after re-incubation. These results suggest that increased CD4 molecules can be induced on the surface of normal human T-cells in vitro. Internalization and accumulation of CD4 molecules occurred in CD4-enriched T-cells with IL-2 pretreatment.

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Internalization of CD4 molecules in human T-cells demonstrated by immuno-electron microscopy

Wang

Lee³

et al. 1992

Histochemistry

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Immunocytochemical demonstration of the binding and internalization of growth hormone in GERL of Chang hepatoma cells

1990

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The binding and internalization of endogenous growth hormone in Chang hepatoma cells were localized on the cell surface and in the Golgi-endoplasmic reticulum-lysosome (GERL) area by various indirect immunocytochemical labeling techniques, namely, peroxidase or colloidal gold conjugated to secondary antibody, and avidin-biotin complex methods. Rabbit antiserum and monoclonal antibodies raised against HPLC-purified porcine growth hormone were used in this study. In fixed material, antigen-antibody complexes were found to be homogeneously distributed along the cell membrane. Control groups showed negative binding on the cell surface. Trypsin treatment before immunolabeling removed antibody binding completely, but hyaluronidase was ineffective. Pretreatment of lectins did not block the recognition of primary antibody to antigen molecules on cell surface. Internalization of the antigen-antibody peroxidase or gold complexes was demonstrated in the cells, which were immunolabeled at 4 degrees C, and then reincubated for 0-30 min at 37 degrees C before fixation. After reincubation, the internalized ligand complexes were found in vesicles near the cell surface or in the GERL area near the Golgi apparatus which, however, did not label for peroxidase. These findings suggest that the trypsin-sensitive growth hormone, specifically bound and internalized into Chang hepatoma cells, is localized in the GERL instead of the Golgi apparatus and might be involved in the mechanism of tumor cell growth.

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Immunocytochemical demonstration of the binding of growth-related polypeptide hormones on chick embryonic tissues

Wang

1989

Histochemistry

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The presence of endogenous growth-related polypeptide hormones, such as growth hormone (GH), somatomadin-C/insulin-like growth factor-1 (SM-C/IGF-1), prolactin (PRL) and Mullerian inhibiting substance (MIS) on chick embryonic tissues have been detected by electron microscopic (EM) immunocytochemistry. Antiserum against GH, anti-SM-C/IGF-1, anti-PRL and anti-MIS were used respectively as primary antibodies for immunolabeling probes by peroxidase (PO) and avidin-biotin complex (ABC)-gold ligands. Cross-reaction studies by ELISA showed negative or weak antigen-antibody interactions. Chick embryos, gonads, and Mullerian ducts (Mds) of various ages were fixed in 2.5% glutaraldehyde for 30 min. Washes in phosphate buffer were administered between each of the following incubations: (i) 2% BSA; (ii) primary antibody; (iii) biotinylated or PO-conjugated secondary antibody; (iv) avidin conjugated with gold particles. SM-C/IGF-1 bindings were negative on 1d embryonic disc, heavily stained on 2d endoderm. However, the GH bindings were found on the embryonic layers of 1d and 2d embryos, and increasing on the luminal epithelial cells of Mds during development. PRL was found in parallel with GH, but in less amount. The 10d Mds were double labeled with anti-SM-C/IGF-1-gold and anti-MIS-PO (MIS-PO), and the results showed SM-C/IGF-I negative, but MIS-PO positive bindings. This study provides the first immunocytochemical evidences for: (i) The presence of GH, SM-C/IGF-1, PRL and MIS bindings on chick embryonic tissues, and further supports their potential role as growth mediators during embryonic development. (ii) The amount of GH and MIS bindings were found correspondingly to their physiological status of Md growth or regression.(ABSTRACT TRUNCATED AT 250 WORDS)

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Immunocytochemical demonstration of the binding and internalization of growth hormone, B-cell stimulating factor-1 and thy-1.2 in murine lymphocytes.

Cited by 3 publications

References 28 publications

Internalization of CD4 molecules in human T-cells demonstrated by immuno-electron microscopy

Internalization of CD4 molecules in human T-cells demonstrated by immuno-electron microscopy

Immunocytochemical demonstration of the binding and internalization of growth hormone in GERL of Chang hepatoma cells

Immunocytochemical demonstration of the binding of growth-related polypeptide hormones on chick embryonic tissues

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