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目的:制备并鉴定小鼠肝炎病毒(MHV)N蛋白的单克隆抗体(mAb)。方法:以Bac-to-Bac杆状病毒表达系统表达的重组N蛋白为免疫原,免疫BALB/c小鼠,应用常规杂交瘤技术将免疫小鼠脾细胞与Sp2/0骨髓瘤进行融合,用间接ELISA方法筛选分泌N蛋白mAb的杂交瘤细胞系。并用Western blot和IFA方法对所获得的mAb进行鉴定。结果:共获得4株稳定分泌抗N蛋白mAb的杂交瘤细胞系,其亚类鉴定2株为IgG2a,1株为IgG2b,1株为IgG1,轻链均为κ型。经鉴定4株mAb均能与重组N蛋白发生特异性反应。结论:成功获得了特异性抗MHV N蛋白的mAb,为进一步研究N蛋白的结构功能及建立诊断学方法奠定了基础。
OBJECTIVE: To prepare and identify monoclonal antibodies (mAbs) for mouse hepatitis virus (MHV) N protein. Methods: BALB / c mice were immunized with recombinant N protein expressed by Bac-to-Bac baculovirus expression system. The spleen cells of immunized mice were fused with Sp2 / 0 myeloma using conventional hybridoma technique. Indirect ELISA for screening hybridoma cell lines secreting N protein mAbs. The obtained mAb was identified by Western blot and IFA. Results: Four hybridoma cell lines stably secreting anti-N protein mAb were obtained. Two strains of IgG2a were identified as subclass, one was IgG2b, the other was IgG1. Four mAbs were identified that reacted specifically with recombinant N protein. CONCLUSION: The mAb with specific anti-MHV N protein has been successfully obtained, which lays the foundation for further study on the structural function of N protein and the establishment of diagnostic methods.