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目的以西尼罗病毒非结构蛋白(nonstructural protein1,NS1)为免疫原制备西尼罗病毒NS1蛋白特异性单克隆抗体,并鉴定其特异性。方法在表达具有良好抗原性的重组西尼罗病毒NS1蛋白基础上免疫Balb/c小鼠,取免疫小鼠的脾细胞与小鼠骨髓瘤细胞进行融合,用间接ELISA法筛选阳性的杂交瘤细胞,并结合免疫荧光(IFA)和免疫印迹对所获得单克隆抗体的特异性进行鉴定,通过竞争抑制试验对m Ab识别的抗原位点进行分析。结果获得39株特异性针对西尼罗病毒NS1蛋白的单克隆抗体,Ig亚类测定结果Ig G3和Ig G2a单抗各2株,Ig G2b单抗5株,Ig G单抗1株,另外29株均为Ig G1。结论成功获得了特异性针对西尼罗病毒NS1蛋白的单克隆抗体,将为进一步建立西尼罗病毒NS1抗原检测方法及探讨NS1蛋白及抗体在西尼罗病毒发病机制中提供依据。
Objective To construct the specific monoclonal antibody of West Nile virus NS1 protein with nonstructural protein1 (NS1) as immunogen and identify its specificity. Methods Balb / c mice were immunized with recombinant West Nile virus NS1 protein with good antigenicity. The spleen cells of immunized mice were fused with mouse myeloma cells. The positive hybridoma cells were screened by indirect ELISA , And the specificity of the obtained monoclonal antibodies was identified by immunofluorescence (IFA) and immunoblotting. Antigen sites recognized by m Ab were analyzed by competitive inhibition assay. Results 39 monoclonal antibodies specific for West Nile virus NS1 protein were obtained. The results of Ig subclass assay showed that Ig G3 and Ig G2a McAb were 2 each, Ig G2b McAb was 5 and Ig G McAb was 1. Strains are Ig G1. Conclusion The monoclonal antibody against NS1 of West Nile virus was successfully obtained, which will provide a basis for further establishing the detection method of NS1 antigen of West Nile virus and exploring the NS1 protein and antibody in the pathogenesis of West Nile virus.