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抗血管紧张素原多克隆抗血清的制备及鉴定

孟庆军, 李剑, 张文红, 张乐宁, 张立藩

第四军医大学航空航天生理学教研室，生物化学与分子生物学教研室，西安710032

摘要

为深入研究血管紧张素原(angiotensinogen , AGT)的表达和组织分布特征，我们以原核表达的AGT蛋白C末端片段为免疫原免疫家兔，制备了针对AGT的多克隆抗血清，并对其进行了ELISA. Western印迹分析及免疫组化鉴定。结果发现，经多次免疫后获得的多克隆抗血清不仅效价高(125 600)，而且能特异性地针对组织内源性及原核表达的AGT蛋白，同时在人和大鼠组织AGT之间会发生交叉反应。以此抗血清进行免疫组化染色，观察到人
胰腺的胰岛细胞及肝内胆管上皮细胞胞浆均有AGT蛋白表达。以上结果提示，利用大肠杆菌融合表达的AGT蛋白可有效刺激家兔产生AGT抗体。本工作为进一步研究AGT乃至局部RAS的生理、病理意义提供了重要的实验工具。

关键词： 血管紧张素原; 原核表达; 抗血清; 特异性

Preparation and inderntification of polyclonal antiserum against arngiotensi noggin

MENG Qing- Jun, Li Jian, ZHANG Wen Hong, ZHANG Le-Ning, ZHANG Li Fan

Department of Aerospace Physiology and Department of Biochemistry and Molecular Biology, the fourth Military Medical University, Xi'an 710032

Abstract

For studying the expression and distribution of angiotensinogen(AGT)，the C to minus of rat AGT gene was expressed in E. coli .Rabbits were immunized with expressed AGT protein and sera from different rabbits were raised.ELISAshowed a high titre(125 600) of the antiserum.With the antise ru m，Western blotting recognized not only the prokaryotic expressed AGT，but also the endogenous AGT protein in liver tissue of both rats and humans .Using this antiserum，i m munohistoche mistry showed the expression of AGT protein in islet cells of human pancreas as well as in epithelium of human
bile duct .These results suggest that the prokaryotic expressed AGT protein is an effective immunogen for the preparation of anti-AGT antiserum .Our present work provides an important tool for study of the path ophysiological role of AGT as well as local renixr angiotensin system.

Key words: angiotensinogen;prokaryotic expression;antiserum;specificity

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引用本文：

孟庆军, 李剑, 张文红, 张乐宁, 张立藩. 抗血管紧张素原多克隆抗血清的制备及鉴定[J]. 生理学报 2003; 55 (1): .

MENG Qing- Jun, Li Jian, ZHANG Wen Hong, ZHANG Le-Ning, ZHANG Li Fan. Preparation and inderntification of polyclonal antiserum against arngiotensi noggin. Acta Physiol Sin 2003; 55 (1): (in Chinese with English abstract).