流行性乙型脑炎病毒E蛋白主要抗原域的原核表达与间接ELISA检测方法的初步建立

Prokaryotic Expression and Establishment of a Putative Indirect ELISAAssay for the MainAntigenic Region of Japanese Encephalitis Virus’s E Protein

重组质粒pET-E转化宿主菌BL21,经1.0mmol/LIPTG诱导,外源基因以包含体的形式获得高效表达。通过Westernblotting检测证明表达产物具有良好的抗原性。以纯化后表达产物作为诊断抗原包被酶标板建立了检测JEV抗体的间接ELISA方法。结果表明,抗原的最佳稀释度为1:2000,血清的最佳稀释度为1:200,待检血清阳性标准初步定为:OD490nm>1.2,且待检血清与阴性血清的OD490nm比值大于2。

The recombinant pET-E plasmid, which includes the gene fragment of the main antigenic region of Japanese encephalitis virus (JEV)’s E protein, was transformed into E.coli BL21, and expressed in very high level as inclusion body after induced with 1.0mmol/L IPTG. The indirect ELISA for detecting JEV E protein antibodies was established after the antigenicity of the recombinant protein was proved by Western-blotting. The optional working circumstances for the ELISA (antigen concentration 1:2000; serum dilution 1:200) were tried out with chess titration. The tested serum of positive criterion of this ELISA is OD490nm>1.2, OD the tested serum/ODthe negative serum >2.1