Zhou Jie, Zhao Li, Hu Jian-Hua, Gao Cheng
Shanghai Research Center of Laboratory Animal, Chinese Academy of Science, Shanghai, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2012 Aug;28(8):841-3.
To prepare and characterize the monoclonal antibodies (mAbs) against N protein of mouse hepatitis virus (MHV).
BALB/c mice were immunostimulated with the purified recombinant N protein expressed by Bac-to-Bac baculovirus expression system. Then the spleen cells of the immunized mice were fused with Sp2/0 myeloma cells by conventional hybridoma techniques. An indirect ELISA using recombinant N protein as antigen was established to screen antibody-producing hybridoma cell lines. Western blotting and IFA were applied to characterize the mAbs.
We obtained four hybridoma cell lines secreting mAbs against recombinant N protein. The subclasses of two mAbs were IgG2a, one IgG2b and the other IgG1, and light chains were both Kappa. Western blotting showed that the mAbs specifically recognized certain antigenic epitope of N protein of MHV.
mAbs against N protein of MHV with high activity and specificity were prepared successfully, which provides a potential basis for further researches on N protein function and diagnostic methods.
制备并鉴定抗小鼠肝炎病毒（MHV）N蛋白的单克隆抗体（mAb）。
用Bac-to-Bac杆状病毒表达系统表达的纯化重组N蛋白免疫刺激BALB/c小鼠。然后通过常规杂交瘤技术将免疫小鼠的脾细胞与Sp2/0骨髓瘤细胞融合。建立以重组N蛋白为抗原的间接ELISA法筛选产生抗体的杂交瘤细胞系。应用蛋白质免疫印迹法（Western blotting）和免疫荧光分析法（IFA）对单克隆抗体进行鉴定。
获得了4株分泌抗重组N蛋白单克隆抗体的杂交瘤细胞系。其中两株单克隆抗体的亚类为IgG2a，一株为IgG2b，另一株为IgG1，轻链均为κ链。蛋白质免疫印迹法显示，这些单克隆抗体能特异性识别MHV N蛋白的某些抗原表位。
成功制备了具有高活性和特异性的抗MHV N蛋白单克隆抗体，为进一步研究N蛋白功能及诊断方法提供了潜在基础。
