单株抗人肝脏微粒体蛋白及细胞色素P450单克隆抗体的制备及初步鉴定

目的:制备抗正常人肝脏微粒体(HLM)及细胞色素P450蛋白的单克隆抗体.方法:采用人肝脏微粒体蛋白作为免疫原免疫Balb/C小鼠,按常规方法融合、克隆,经间接ELISA法筛选抗体阳性杂交瘤细胞株,通过单抗亚类、Western blot对抗体进行初步鉴定.结果:成功地制备了一株抗肝微粒体蛋白的单克隆抗体(MAb),命名为MAb 10C7.用鼠单抗亚类快速鉴定纸条鉴定10C7为IgG2b.Western blot分析显示,MAb 10C7可特异识别相对分子量为46kD的人肝脏微粒体蛋白.结论:MAb 10C7抗正常人肝脏微粒或细胞色素P450蛋白的特异性强,对相关蛋白质的功能研究有较好的应用前景.

Preparation and Preliminary Identification of HLM and CYP450 Monoclonal Antibody

Objective:The present study was intended to prepare a kind of specific MAb against HLM protein by cell engineer method and to identify its immunological characteristics.Methods:The Balb/C mouse was immunized with mixed HLM proteins 3 to 4 times.And it was boosted with HLM proteins three days before the hybridization.The spleen cells of immunized mouse were hybridized with SP2/0 cells with 50% PEG 4 000.The hybridized cells were cloned ac- cording to standard method.The MAb was identified by ELISA,and Western Blot.Results:The mouse was immunized successfully using the mixed HLM proteins.An antibody-secreting hybridoma cell line,10C7,was obtained.The isotype of the MAbs 10C7 was IgG2b tested by mouse monoclonal antibody isotyping kit.Immumodominant antigen bind- ing of MAb 10C7 was located at molecular weight about 46kD via Western Blot.Conclusion:One hybridoma cell line secreting MAb against HLM protein was obtained.The MAb 10C7 might be useful in HLM immunoassay.