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目的采用大肠杆菌系统表达乙脑病毒NS1蛋白,并对表达产物进行免疫原性分析。方法将乙脑病毒NS1基因克隆入pET30a载体中,表达产物经Ni+亲和层析纯化后行SDS-PAGE和Western blot分析;并将纯化产物免疫新西兰大白兔,获得的免疫血清作间接ELISA及免疫荧光检测。结果重组JEV NS1蛋白获得表达并纯化成功;Western blot结果显示纯化蛋白能与JEV免疫的鼠多克隆腹水反应;兔免疫血清的间接ELISA结果显示血清效价达到1∶51 200以上;免疫荧光结果显示血清作1∶50、1∶100、1∶200稀释时,JEV感染的BHK细胞均显示荧光。结论重组NS1蛋白能在大肠杆菌系统中获得表达;纯化产物具有免疫原性,为进一步研究NS1蛋白的生物学特性及乙脑病毒的诊断试剂提供参考依据。
Objective To express the encephalitis virus NS1 protein in Escherichia coli and to analyze the immunogenicity of the expressed product. Methods The NS1 gene of JE virus was cloned into pET30a vector. The expressed product was purified by Ni + affinity chromatography and analyzed by SDS-PAGE and Western blot. The purified product was immunized with New Zealand White rabbits and the obtained serum was used for indirect ELISA and immunization Fluorescence detection. Results The recombinant JEV NS1 protein was expressed and purified successfully. The result of Western blot showed that the purified protein reacted with polyclonal ascites of JEV-immunized mice. The result of indirect ELISA showed that the serum titer reached 1:51 200 and the result of immunofluorescence The BHK cells infected with JEV showed fluorescence when the serum was diluted 1:50, 1: 100 and 1: 200. Conclusion The recombinant NS1 protein can be expressed in Escherichia coli system. The purified product has immunogenicity and provides a reference for further study on the biological characteristics of NS1 protein and diagnostic reagent for JE virus.