乙型脑炎病毒(JEV)单克隆抗体的制备及初步应用

目的制备乙型脑炎病毒(JEV)单克隆抗体,应用于猪源性生物制品中JEV的检测。方法提纯病毒抗原,免疫BALB/c小鼠,取脾细胞与SP2/0骨髓瘤细胞融合,经间接ELISA方法筛选和有限稀释法三次克隆获得4株杂交瘤细胞,并应用琼脂扩散、酶联免疫吸附试验和免疫荧光方法进行抗体鉴定和样品检测。结果获得4株稳定分泌JEV单克隆抗体的杂交瘤细胞株G10、F4、C10、A5,免疫荧光抗体实验证实是特异的抗JEV抗体,其免疫球蛋白亚类G10和F4为IgG1,A5为IgM,C10未鉴定出其亚类。G10和F4腹水效价为105以上,C10和A5腹水效价为102以上。结论成功制备了抗JEV单克隆抗体,建立了以单抗介导的间接ELISA和间接IFA检测方法,并应用于猪源性生物制品中JEV的检测。

Preparation and Preliminary Application of Monoclonal Antibodies Against JEV

Objective\ To prepare Monoclonal Antibodies against JEV and apply preliminary the antibodies to the detection.Methods\ BALB/c mice were immunized with purified Japanese Encephalitis Virus(JEV).The immunized spleen cells of these mice were fusing with SP2/0 myeloma cells.Four hybridoma secreting monoclonal antibodies(McAbs) against JEV were prepared after screening by ELISA and cloning three times by limiting dilution.Results\ Four hybridomas were obtained,named A5,Gl0,F4,and C10 respectively,which were able to secrete McAbs stably.The immunoglobulin subclass of G10 and F4 were IgG1 and A5 was IgM,the subclass of C10 was not identified;the ELISA titers of C10 and F4 ascites were above 10~5,while the ELISA and IFA titers of C10 and A5 ascites were above 10~2.Conclusion\ Monoclonal Antibodies against JEV were successfully prepared,the method of IFA and ELISA based on McAb was developed for the detection of JEV in the biological products from the swine for human use.