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首页> 外文期刊>Life sciences >Generation of polyclonal antiserum against the growth hormone secretagogue receptor (GHS-R): evidence that the GHS-R exists in the hypothalamus, pituitary and stomach of rats.
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Generation of polyclonal antiserum against the growth hormone secretagogue receptor (GHS-R): evidence that the GHS-R exists in the hypothalamus, pituitary and stomach of rats.

机译：对抗生长激素促分泌素受体（GHS-R）的多克隆抗血清的产生：证据表明GHS-R存在于大鼠的下丘脑，垂体和胃中。

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摘要

Growth hormone (GH) secretagogues (GHSs), which stimulate GH secretion, are synthetic compounds that act through the GHS receptor (GHS-R) which has been recently cloned. We raised an antiserum in a rabbit against a synthetic peptide corresponding to amino acid residues 248-260 of the third intracellular loop of the rat GHS-R. A competitive immunoassay showed that the antiserum had a specific affinity for the target peptide. To confirm the specificity of the antiserum, the GHS-R cDNA was stably expressed in COS-7 cells. In Western blot analysis, the band was detected at 44 kDa in the extracts from COS-7 cells expressing GHS-R (COS-7/tf3-2) but not in those from wild-type COS-7 cells. Furthermore, while COS-7/tf3-2 cells were strongly immunostained for GHS-R, no GHS-R-like immunoreactivity was observed in wild-type COS-7 cells. Immunoreactive bands were also observed at approximately 46 kDa in the extracts from rat hypothalamus, pituitary and stomach by Western blot analysis. These studies are the first to show the existence of GHS-R protein in the stomach. The antiserum for the GHS-R is sensitive and specific, and it would be useful for clarifying the roles of GHS/ghrelin.
机译：刺激GH分泌的生长激素（GH）促分泌素（GHS）是通过最近被克隆的GHS受体（GHS-R）起作用的合成化合物。我们在兔体内产生了针对合成肽的抗血清，该合成肽对应于大鼠GHS-R第三细胞内环的氨基酸残基248-260。竞争性免疫测定表明抗血清对靶标肽具有特异性亲和力。为了证实抗血清的特异性，GHS-R cDNA在COS-7细胞中稳定表达。在蛋白质印迹分析中，在表达GHS-R（COS-7 / tf3-2）的COS-7细胞的提取物中以44 kDa检测到该条带，而在野生型COS-7细胞的提取物中未检测到该条带。此外，尽管对COS-7 / tf3-2细胞的GHS-R进行了强力免疫染色，但在野生型COS-7细胞中未观察到GHS-R样的免疫反应性。通过Western印迹分析在大鼠下丘脑，垂体和胃的提取物中还观察到约46kDa的免疫反应带。这些研究首次表明胃中存在GHS-R蛋白。 GHS-R的抗血清具有敏感性和特异性，有助于阐明GHS / ghrelin的作用。

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