HIV-1囊膜糖蛋白gp41三聚体结构域基因的表达及其单克隆抗体的制备

Expression of a trimeric structural domain of the HIV-1 transmembrane glycoprotein and preparation of monoclonal antibodies

目的 :表达HIV 1囊膜糖蛋白gp4 1三聚体结构域基因N5 1(L6 )C4 6融合蛋白 ,并制备针对该融合蛋白的单克隆抗体 (mAb) ,为筛选抗HIV多肽及分析gp4 1表位的免疫原提供抗体工具。方法 :在BL2 1(DE3)中诱导表达N5 1(L6 )C4 6融合蛋白 ,经mAbNC 1鉴定后 ,采用小鼠腹股沟皮下NC膜免疫的方法免疫小鼠 ,并进行细胞融合、克隆化制备抗N5 1(L6 )C4 6mAb ,用ELISA法初步鉴定其特异性位点。结果 :获得了高表达的N5 1(L6 )C4 6融合蛋白 ,并能与识别特异性空间构象的mAbNC 1反应。用该融合蛋白免疫小鼠后 ,获得了 6株抗N5 1(L6 )C4 6的mAb ,这 6株mAb均特异识别兔抗N36 (L6 )C34抗体捕获的融合蛋白 ,但不与N36或C34多肽片段反应。结论 :得到高效表达融合蛋白N5 1(L6 )C4 6 ,并呈现gp4 1核心结构空间构象。用此蛋白免疫小鼠得到 6株特异结合gp4

Objective:To express N51(L6)C46 gene of HIV 1 gp41 trimeric structural domain in E.coli BL21(DE3) and generate monoclonal antibodies against N51(L6)C46.Methods:N51(L6)C46 was induced in E.coli BL21(DE3).The expressed protein was detected with the antibody NC 1.McAb clones against N51(L6)C46 were prepared by hybridoma technique and identified by ELISA.Results:N51(L6)C46 was induced successfully and the expressed protein was identified by SDS PAGE and detected with NC 1.The six of anti N51(L6)C46 could bind N51(L6)C46,but couldn′t bind N36 or C34.Conclusion:The results suggested the six antibodies may be react with the oligomeric forms of gp41.