定朱宁 王嘉仪 田恩江 汤新之 300070 天津医科大学生化教研室(朱宁、王嘉仪、汤新之)；内分泌研究所(田恩江) 中华内分泌代谢杂志 1998 0 14 6


    关键词：抗磷酸酪氨酸抗体；红细胞；胰岛素受体；自身磷酸化；酶联免疫吸附测定 期刊 zhnfmdxzz 0 论著 fur -->


    

【摘要】 目的 建立人红细胞胰岛素受体自身磷酸化测定法。方法 应用合成的磷酸酪氨酸-钥孔帽贝血蓝蛋白(PY-KLH)免疫家兔得到抗血清，将纯化的抗磷酸酪氨酸抗体作为第一抗体，羊抗兔Ig酶结合物(辣根过氧化物酶标记)作为第二抗体，建立双抗体夹心酶联免疫吸附测定法。结果 批内变异系数为8.4%，批间变异系数为9.0%。20例健康人的红细胞IR的自身磷酸化活性，测定值为0.150±0.022nmol PY/mg受体蛋白。在体外，胰岛素在10^-6 mol/L时对自身磷酸化达到最大激活。结论 用酶免法测定人红细胞胰岛素受体灵敏、特异、简便，为胰岛素作用机制的理论研究、胰岛素受体基因突变的检测和糖尿病及其它胰岛素抵抗综合症的病因及治疗的探讨提供了有效的方法。

ELISA method forautophosphorylation of insulin receptor from human erythrocytes

    Zhu Ning,Wang Jiayi,Tian Enjiang,et al.Department of Biochemistry,Tianjin Medical University,Tianjin,300070

【Abstract】 Objective To establisha method of detecting autophosphorylated insulin receptor from erythrocytes. Methods Phosphotyrosinewas first combined with hemocyanin from keyhole limpets (KLH). Then, rabbits were immunedwith the complete antigen and antisera were gained. The ELISA system consisted of purifiedantiphosphotyrosine antibody as first antibody and sheep antirabbit lgG-HRP as secondantibody. Results CV within the batch was 8.4%. CV between the batcheswas 9.0%. The value for autophosphorylation of insulin receptor from erythrocytes in 20normal individuals was 0.150±0.022nmoL PY/mg receptor protein. In vitro,autophosphorylation of insulin reached maximum activation at 10^-6 mol/L insulin.Conclusion The ELISA method is sensitive, specific and simple forautophosphorylation of insulin receptor. It provides a useful tool for theoreticalresearch of mechanism of insulin action, detection of gene mutation of insulin receptorand exploration of cause and care for insulin resistance.

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