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目的:制备针对西尼罗病毒NS1蛋白的特异性单克隆抗体。方法:以原核表达的重组NS1蛋白为免疫原,免疫BALB/c小鼠并通过常规杂交瘤技术将免疫后小鼠脾细胞与SP2/0细胞进行融合。以真核表达的重组NS1蛋白为检测用抗原,建立间接ELISA检测方法筛选分泌NS1蛋白单克隆抗体的杂交瘤细胞。并利用Western blot和IFA试验对所获得的单克隆抗体进行鉴定。结果:共获得了2株稳定分泌NS1蛋白单克隆抗体的杂交瘤细胞株,分别命名为WN-1C10和WN-3D10,其亚类鉴定分别属于IgG2a和IgG1。这2株单克隆抗体均能与西尼罗病毒NS1蛋白和病毒抗原发生特异性反应,而与日本脑炎病毒无交叉反应。结论:本实验成功制备出针对西尼罗病毒NS1蛋白的特异性单克隆抗体,为我国建立西尼罗病毒与日本脑炎病毒的血清学鉴别诊断方法奠定了基础。
Objective: To prepare specific monoclonal antibodies against West Nile virus NS1 protein. Methods: BALB / c mice were immunized with recombinant NS1 protein expressed in prokaryotic cells and splenocytes of immunized mice were fused with SP2 / 0 cells by conventional hybridoma technique. The eukaryotic recombinant NS1 protein was used as detection antigen, and an indirect ELISA method was established for screening hybridoma cells secreting monoclonal antibody against NS1 protein. The obtained monoclonal antibodies were identified by Western blot and IFA assay. Results: Two hybridoma cell lines stably secreting monoclonal antibodies against NS1 protein were obtained and named as WN-1C10 and WN-3D10, respectively. The subclasses were identified as IgG2a and IgG1. Both of these monoclonal antibodies reacted specifically with the NS1 protein of West Nile virus and the viral antigens but did not cross-react with the Japanese encephalitis virus. Conclusion: The specific monoclonal antibody against NS1 protein of West Nile virus was successfully prepared in this study, which laid the foundation for the differential diagnosis of West Nile virus and Japanese encephalitis virus in China.