丙型肝炎病毒单克隆抗体的制备及其鉴定

Establishment and Identification of Monoclonal Antibody of Hepatitis C Virus

目的押以基因工程表达的丙型肝炎病毒抗原蛋白(HCAg)为抗原,建立分泌单克隆抗体(McAb)的杂交瘤细胞系,并对其分泌的McAb进行鉴定。方法:用纯化HCV基因工程表达抗原(HCAg)免疫Balb/c小鼠,取脾细胞及骨髓瘤细胞按常规方法融合、筛选、克隆化及腹水注射制备McAb。采用ELISA及分片断鉴定技术进行鉴定。结果押筛选出5株能稳定分泌特异性抗-HC的McAb杂交瘤细胞株,经多次复苏传代仍能稳定分泌抗体,特异性强,效价高。与不同抗原蛋白的ELISA反应呈特异性阳性反应。分片断鉴定结果显示,所获5株McAb主要为抗HCV核心蛋白和非结构蛋白NS3。结论:此5株丙肝杂交瘤细胞株分泌的抗体对丙肝抗原具有特异亲和性,为丙型肝炎病毒抗原诊断试剂的研制提供了关键技术基础。

Objective： To establish hybridoma cell line secreting monoclonal antibody （McAb） by hepatitis C virus antigen（HCAg） and identify the McAb. Methods： Balb/c mice were immunized with HCAg. The spleen cells and marrow tumor cells were regularly fused, screened, cloned and ascites injected for McAb preparing. The specificity was identified by ELISA and line immunoassay. Results： Five hybridoma cell lines stably secreting specific McAb against HCAg were screened, McAb of the five hybridoma cell lines kept stably secreting after several times of resuscitation and regeneration, and perfect specificity and higher efficiency were established. It was positively related to different antigen protein ELISA reaction. The main components of the five McAb were anti-core and anti-NS3 by line immunoassay. Conclusion： The anti-HC monoclonal antibody secreted by the five hybridoma cell lines showed perfect specificity and strong affinity with HCAg, and it provides a foundation for clinical early diagnosis and therapy research.