重组H5亚型流感病毒HA1抗原的制备及其特性分析

Characterization of the E.coli-expressed HA1 protein of H5 subtype influenza virus

为获得重组表达的H5亚型流感病毒血凝素(HA1)抗原,将PCR扩增的H5亚型流感病毒HA1基因克隆到原核表达载体pET-28a,构建重组原核表达质粒pETHA1-H5。结果显示,转化pETHA1-H5的大肠杆菌BL21(DE3)在IPTG的诱导下以包涵体形式表达了HA1蛋白。HA1重组蛋白能与不同禽类(鸡、鸭和鹅)H5亚型流感病毒阳性血清发生特异性反应,同时以HA1重组蛋白为抗原制备的单克隆抗体具有与H5亚型完整病毒粒子抗原反应的能力,说明重组表达的HA1蛋白保留了自然条件下H5亚型流感病毒的抗原特征。将重组表达的HA1蛋白作为ELISA抗原检测禽血清中的H5亚型流感病毒抗体,以HI试验结果为参考,两者的符合率为96.6%,证明HA1重组蛋白作为ELISA检测抗原是可行的。

To produce the recombinant hemagglutinin（HA1）protein of H5 subtype avian influenza virus（AIV）, the recombinant plasmid pETHA1-H5 was constructed by the RT-PCR-amplified HA1 gene and then cloned into a pET-28a expression vector. The results showed that the insoluble HA1 protein was expressed in the Escherichia coli BL21 （DE3）strain transformed with the plasmid pETHA1-HS, and can be recognized with chicken,duck and goose AIV antisera （H5 subtype）. The monoclonal antibodies to the prokaryotically expressed HA1 protein reacted with the influenza virus antigen（H5 subtype）. The data confirmed that the E. coli-expressed HA1 protein remains the antigenic characteristics of H5 subtype avian influenza virus. In detecting the antibody to H5 subtype AIV,the HA1- based ELISA shares a higher identity to hemagglutination inhibition. The result illuminated that the recombinant HA1 protein has a potential as an ELISA antigen for monitoring antibody to H5 subtype avian influenza virus.