抗人肝微粒体蛋白单克隆抗体细胞系的建立及鉴定

Preparation of monoclonal antibody against human liver microsome proteins

目的初步建立抗人肝微粒体蛋白单克隆抗体规模化制备技术平台,为制备抗人细胞色素P450(CYP450)亚型蛋白的单克隆抗体奠定基础。方法按常规方法进行细胞融合,以酶联免疫吸附测定法(ELISA)筛选杂交瘤,以ELISA、免疫组化(IH)、免疫印迹(Western Blot)、免疫沉淀(IP)对单克隆抗体加以鉴定。结果进行6次融合,共筛出216株杂交瘤,其分泌抗体类型为IgG1I、gG2aI、gG2b、及IgM。免疫组化(IH)图片上,人肝细胞浆内均可见阳性颗粒。免疫沉淀(IP)和免疫印迹(Western Blot)结果显示,所制备抗体与人肝微粒体蛋白有特异性结合。结论筛选制备的单克隆抗体杂交瘤能分泌抗人肝微粒体蛋白单克隆抗体,并且有较强特异性,为下一步研究提供了基础与技术平台。

Objective To establish a technical platform for the preparation and identification of monoclonal antibodies against human liver microsome proteins. Methods Mice were immunised with human liver microsome proteins. Myeloma cells were fused with the spleenocytes. Monoclonal hybridomas were screened by indirect enzyme-linked immunosorbent assay（ELISA） ;and were identified by ELISA, Immunohistochemistry（IH）, Western Blot and Immunoprecipitation（IP）. Results Hybridomas producing monoclonal antibodies were obtained. The subtypes of immunoglobulins produced were identified as IgG1, IgG2a, IgG2b and IgM by immunol- ogy method. Brown particles were seen in normal human liver tissues in IH photos. Conclusion The monoclonal antibodies produced are specific for human hepar microsome proteins.