丙型肝炎病毒囊膜蛋白基因疫苗的构建及其诱导小鼠体液免疫应答

将丙型肝炎病毒（HCV）H77株全长囊膜蛋白基因克隆到真核表达载体pcDNA4.0的CMV启动子下游，采用磷酸钙转染法将重组质粒转入293T细胞，用流式细胞仪检测细胞瞬时表达的目的蛋白。将构建的重组质粒肌注BALB/c小鼠，用表达HCV囊膜蛋白的SP2/0细胞作为抗原，用流式细胞仪检测小鼠血清HCV囊膜蛋白抗体。再用原核系统表达的E2蛋白作为抗原,Western Blot检测免疫小鼠血清抗体。试验结果显示,囊膜蛋白E1E2在293T细胞膜上进行了瞬时表达，基因疫苗免疫小鼠后产生了抗HCV囊膜蛋白的抗体，该抗体能与表达HCV囊膜蛋白的SP2/0细胞特异性结合，Western Blot表明该抗体也能与原核系统表达的E2蛋白结合。

Construction of hepatitis C virus E1E2 envelope protein gene vaccine and its humoral immune response in BALB/c mice

Hepatitis C virus (HCV) H77 strain E1E2 gene vaccine was constructed by inserting full-length cDNA of HCV E1E2 into an eukaryotic expression vector pcDNA4.0.The recombinant plasmid was transfered into eukaryotic cells 293T by calcium phosphate transfection method and transient expressive E1E2 envelope protein was analyzed by FACS.BALB/c mice were injected intramuscularly with the recombinant plasmid.Anti-HCV E1E2 antibody was screened by FACS,with cells SP2/0 which expressed protein HCV E1E2 as antigen.Moreover,the antibody was also analyzed by Western blot and the antigen was E2 protein which was expressed in E.coli.The results showed that protein HCV E1E2 was expressed transciently in cells 293T.Specific anti-E1E2 antibody could be detected in DNA immuned mouse serum by FACS and the antibody could react specially to cells SP2/0 which expressed protein HCV E1E2.Western blot analysis showed that DNA immuned mouse serum could react specially to protein E2 which was expressed by E.coli.