针对人 DNAH2 蛋白的鼠源单克隆抗体的制备

目的 制备特异性的鼠源单克隆抗 DNAH2（Homo sapiens dynein， axonemal， heavy chain 2） 抗体。 方法首先针对人 DNAH2 蛋白 N 端 1~300 aa 序列， 构建可表达 His 标签的免疫原融合表达质粒， 在大肠杆菌内以 IPTG 诱导 His-免疫原的表达并纯化；然后免疫 BALB/c 小鼠， 分离出脾淋巴细胞， 并与骨髓瘤细胞融合， 形成杂交瘤细胞；最后以酶联免疫吸附测定（ELISA）、蛋白免疫印迹法（Western blot）筛选阳性杂交瘤细胞。 结果 利用 IPTG 有效诱导了大肠杆菌内 DNAH2 免疫原的表达； 筛选的阳性杂交瘤细胞检测出 DNAH2 蛋白条带。 结论 成功制备针对人 DNAH2 蛋白的鼠源单克隆抗体。

The preparation of the mouse monoclonal antibodies specific for the DNAH2 protein

Objective To prepare specific mouse monoclonal antibodies against Homo sapiens dynein， axonemal， heavy chain 2 (DNAH2). Methods Firstly, recombinant plasmid encoding His tagged immunogen, targeting N-terminal sequence of DNAH2 protein (1- 300 aa), in E. coli was constructed. IPTG was used to induce the expression of Hisimmunogen, which was then purified and immunized in BALB/c mice. Hybridoma cells were obtained through the fusion between myeloma cells and splenocytes isolated from BALB/c mice. Finally, ELISA and Western blot assays were performed to screen the positive hybridoma. Results IPTG was used efficiently to induce the expression of DNAH2 immunogen in E. coli. DNAH2 protein bands were detected in screened positive hybridoma. Conclusion Mouse monoclonal antiDNAH2 antibody is prepared successfully.