用重组痘苗病毒作载体表达甲型肝炎病毒抗原

EXPRESSION OF HEPATITIS A VIRUS PROTEINS BY RECOMBINANT VACCINIA VIRUS

含有甲型肝炎病毒基因组全部开放读码框架cDNA的重组痘苗病毒,感染143TK^-细胞后,成功地表达了甲肝病毒抗原。ELISA测定其滴度为1:8。免疫荧光法显示,荧光呈颗粒状分布在胞浆内。Western blot证实,重组痘苗病毒表达的甲肝病毒抗原有两条特异性带,一条分子量为29kd,与甲肝病毒抗原VP2相对应;另一条分子量为64kd,可能是未全裂解的甲肝病毒抗原蛋白或聚合体。初步动物免疫实验表明,痘苗病毒表达的甲肝病毒抗原具有较好的免疫原性。

A complete HAV cDNA open reading frame was inserted into a vaccinia virus vector. DNA hybridization confirmed that the HAV cDNA was correctly integrated into vaccinia virus DNA. HAV proteins were successfully expressed in mammalian cells infected with the recombinant viruses. The products had quite good antigenicity in rabbit. They reacted with chimpanzee and rabbit anti-HAV sera in Western blot and IF assay. They also reacted well with anti-HAV McAb. The antigen titer was 1 : 8 by ELISA method. The antigen was located in the cytoplasm of infected cells, with specific granular particles by IF assay. The expressed HAV proteins showed two bands in Western-blot: One protein band was 29kd,the same size as VP2; the other one was much larger, 64kd, which may be a VP1 dimer or a kind of HAV protein that was not cut well by the HAV encoded proteases. Studies on the detailed properties of the expressed HAV proteins are now under way.