口蹄疫病毒受体猪源整联蛋白β6亚基配体结合域单克隆抗体的制备

Preparation of monoclonal antibodies against porcine integrin β6 subunit ligand-binding domain as foot-and-mouth disease virus receptor

以纯化的口蹄疫病毒受体猪源整联蛋白β6亚基配体结合域重组蛋白为抗原,免疫雌性BALB/c小鼠,经3次免疫后取其脾细胞与SP2/0骨髓瘤细胞融合,制备口蹄疫病毒受体猪源整联蛋白β6亚基配体结合域单克隆抗体(McAb);采用有限稀释法和间接ELISA克隆和筛选阳性杂交瘤细胞株,并用SDS-PAGE、间接ELISA以及间接免疫荧光法对制备的McAb的特异性进行鉴定。结果显示,成功获得5株能稳定传代并分泌抗β6亚基配体结合域的杂交瘤细胞株,分别命名为C4C7、E7C7、B8C9、C2D8和B7B6,其分泌的McAb为IgG2b(C2D8)和IgG2a(C4C7、E7C7、B8C9、B7B6)亚类。制备的口蹄疫病毒受体猪源整联蛋白β6亚基配体结合域McAb可为深入研究整联蛋白αvβ6在口蹄疫病毒感染过程中的作用奠定基础。

Monoclonal antibodies（McAbs） C4C7, E7C7, B8C9,C2D8 and B7B6 against porcine integrin β6 subunit ligand-binding domain（LBD） as foot-and-mouth disease virus（FMDV） receptor were produced by fusing SP2/0 myeloma cells with spleen cells of female BALB/c mice immunized with purified porcinederived integrin β6 subunit LBD recombinant fusion protein as antigen for 3 times. Masculine hybridoma cell was cloned and sieved by limiting dilution assay and indirect enzyme-linked immunosorbent assay （iELISA）. McAbs specifically reacted with integrin β6 subunit LBD recombinant fusion protein were identified by SDS-PAGE, iELISA and indirect immunofluorescence assay （IFA）. Five stable hybridoma cell strains, named C4C7,E7C7, B8C9, C2D8 and B7B6, were established. These McAbs were secreted as IgG2a （C4C7,E7CT,BSC9 and B7B6） and IgG2b （C2D8） subtypes,respectively. The prepared McAbs could provide foundation for studying the roles of integrin αvβ6 in FMDV infection.