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目的:以人高致病性禽流感H5N1病毒M2蛋白为靶抗原,制备针对H5N1病毒M2蛋白的单克隆抗体,并对制备的单克隆抗体进行鉴定。方法:用针对H5N1病毒M2蛋白胞外区设计的四分肽作为免疫原免疫BALB/c小鼠,将免疫后的脾细胞与SP2/0骨髓瘤细胞融合,用间接ELISA方法筛选出阳性细胞,经多次亚克隆筛选出稳定分泌特异性抗体的杂交瘤细胞株,制备单抗腹水并进行抗体效价测定及亚类鉴定。结果:获得10株能稳定分泌抗H5N1病毒M2蛋白单克隆抗体杂交瘤细胞株,抗体效价高,抗原特异性强。结论:成功地获得了针对H5N1病毒M2蛋白的单克隆抗体,为H5N1病毒感染的诊断、治疗及病毒致病机制研究奠定了基础。
OBJECTIVE: To prepare the monoclonal antibody against M2 protein of H5N1 virus using M2 protein of human H5N1 virus as target antigen, and to identify the prepared monoclonal antibody. Methods: BALB / c mice were immunized with tetrapeptide designed for extracellular domain of H5N1 virus M2 protein. The immunized spleen cells were fused with SP2 / 0 myeloma cells, and the positive cells were screened by indirect ELISA. After multiple subcloning screening of hybridoma cell lines secreting specific antibodies, monoclonal antibody ascites was prepared for antibody titer determination and subclass identification. Results: Ten hybridoma cell lines that can stably secrete M2 monoclonal antibody against H5N1 virus were obtained. The antibody titer is high and the antigen specificity is strong. Conclusion: The monoclonal antibody against M2 protein of H5N1 virus has been successfully obtained, which lays the foundation for the diagnosis and treatment of H5N1 virus infection and the pathogenesis of virus.