T7噬菌体衣壳蛋白P11的原核表达、纯化及其单克隆抗体的制备  

Expression and purification of T7 bacteriophage capsid protein P11 and preparation of monoclonal antibody against P11

作　　者：陈亮[1] 高云英[1] 王华茂[2] 徐蓉[2] 刘素霞[2] 李宗海[2] 

机构地区：[1]西北农林科技大学动物医学院,陕西杨凌712100 [2]上海市肿瘤研究所,癌基因及相关基因国家重点实验室,上海200032

出　　处：《细胞与分子免疫学杂志》2008年第5期482-483,490,共3页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家重点基础研究发展计划(973)资助项目(2004CB518802)

摘　　要：目的:表达、纯化T7噬菌体衣壳蛋白P11,并制备其单克隆抗体(mAb)。方法:克隆并表达T7噬菌体P11蛋白,其氨基端带有6-His标签。纯化后的蛋白免疫BALB/c小鼠,经融合、筛选制备特异性mAb。结果:成功表达了P11蛋白。SDS-PAGE显示所表达蛋白的相对分子质量(Mr)约为27000。获得了1株稳定分泌抗P11抗体的杂交瘤细胞株(2G11),其分泌的mAb的Ig亚类(型)为IgG2b。ELISA检测,对应腹水mAb的效价为1∶8.1×105。Western blot结果显示抗P11mAb具有良好的特异性。结论:成功地制备了P11蛋白及其mAb。AIM： To express capsid tail protein PII of T7 bacteriophage and produce mouse monoclonal antibody （mAb） against the protein. METHODS： P11 protein was cloned and recombinant P11 protein was expressed as a fusion protein with an N-terminal 6-His tag. The purified protein was used to immunize BALB/c mouse. The specificity of mAb was analyzed by ELISA and Western blot. RESULTS： P11 protein was successfully expressed and pu- rified. SDS-PAGE analysis showed that the molecular weight of the expressed protein was approximately 27 kd. One hybridoma cell （2Gll） secreting mAb against P11 was developed. The isotype of the mAb was IgG2b. ELISA detection showed that titers of mAb was 1：8.1 × 10^5 in ascites. Western blot analysis proved mAb obtained could react specifically to the recombinant pll protein. CONCLUSION： Recombinant P11 protein and mAbs were successfully prepared.

关 键 词：T7噬菌体 P11蛋白 原核表达 单克隆抗体 

分 类 号：R392.11[医药卫生—免疫学]