重组人促甲状腺激素β亚基的切胶纯化及多克隆抗体制备  

Gel purified recombinant protein as immunogen to generate polyclonal antibodies against human thyrotropin β subunit(hTSHβ)

作　　者：李敬华[1] 柳明波[1] 马彦[1] 左爱军[1] 孙蓓(指导)[1] 梁东春[1] 郭刚[1] 

机构地区：[1]天津医科大学代谢病医院/内分泌研究所,卫生部及天津市激素与发育重点实验室,天津300070

出　　处：《中国免疫学杂志》2010年第5期428-431,共4页Chinese Journal of Immunology

基　　金：天津市“重中之重”学科建设专项基金资助项目(05YFGDSF02700)

摘　　要：目的:切胶纯化重组人促甲状腺激素亚基(hTSHβ),免疫家兔制备多克隆抗体,为人TSH测定试剂盒的国产化奠定基础。方法:将含有重组人促甲状腺激素β亚基(hTSHβ)的大肠杆菌表达产物进行SDS-PAGE分离,切取相应分子量的蛋白条带,研磨回收后用于免疫新西兰大耳朵白兔,ELISA监测血清中抗hTSHβ抗体的效价,饱和硫酸铵沉淀及亲和层析自血清中纯化抗体。结果:切胶后获得了高纯度的重组GST-hTSHβ蛋白,所免疫家兔产生了针对hTSHβ的多克隆抗体,效价可达1:12000。结论:切胶纯化的重组hTSHβ蛋白可成功诱导兔产生高效价的抗hTSHβ多克隆抗体。Objective：To prepare polyclonal antibody against human thyrotropin β subunit with recombinant human TSHβ,expressed in Escherichia coli and purified from the PAGE gel,as the immunogen.Methods：Raw E.coli expressions of rhTSHβ were subjected to SDS-PAGE,gel pieces containing proteins corresponding to the molecular weight of GST-rhTSHβ were excised and homogenated in liquid nitrogen.Extracted protein was dissolved in PBS solution and used as immunogen to immune rabbits after being verified as GST-fusioned protein by western blot.The titter of the anti-hTSHβ antiserum was monitered by ELISA.The antiserum was collected after the titter reach 1：12 000 and then IgG was purified by chromatography followed by ammonium sulfate precipitation.Results：Highly purified GST-rhTSHβ protein was obtained from the gel,which could successfully induce the generation of high-tittered anti-hTSHβ antibody in the rabbits.Conclusion：Above mentioned procedure applies us an option for preparing recombinant proteins as the immunogen and generating antibodies from the rabbit.

关 键 词：人促甲状腺激素 免疫 人促甲状腺激素抗体 重组蛋白 

分 类 号：R392.6[医药卫生—免疫学]