基于基因免疫制备抗人绒毛膜促性腺激素β亚基单克隆抗体及其特性研究

Preparation of the anti-human chorionic gonadotrophin β-subunit monoclonal antibodies based on the genetic immunization with recombinant plasmid and characterization of its biological properties

用真核表达人绒毛膜促性腺激素β亚基((human chorionic gonadotrophinβ,hCGβ)的重组质TR421-hCGβ免疫BALB/c小鼠,采用B淋巴细胞杂交瘤技术进行细胞融合,ELISA法筛选阳性细胞株,经过多次的克隆化培养,最终获得10株持续、稳定分泌抗hCGβ单克隆抗体的杂交瘤细胞株。随机抽取6H1杂交瘤细胞进行抗体的制备、纯化及免疫学特性的分析。间接ELISA法证明6H1单抗属于IgG2a亚类。Western blot证明6H1单克隆抗体可以特异性结合hCGβ,间接免疫荧光和流式细胞仪等结果表明,6H1单克隆抗体能够不同程度的结合不同来源的肿瘤细胞膜上表达的hCGβ分子,为进一步研究其生物学功能奠定了物质基础。

The eukaryotic expression vector TR321 hCGβ encoding the β subunit of human chorionic gonadotrophin （hCGβ） was used to immunize BALB/c for the prepration of corresponding monoclonal antibodies. By means of the routine hybridoma technique for B lymphocytes of cell fusion, repeated screening by ELISA and multiple cell cloning, 10 hybridoma cell lines, capable of secreting anti-hCGβ monoclonal antibodies continously, were thus obtained, in which 6H1 hybridoma was random ly selected to prepare the specific antibody, to purify the secreted antibodies and to analyse its immunologic properties. As demonstrated by indirect ELISA assay, the 6H1 monoclonal antibody was isotyped to be IgG2a. In Western blot assay it was demonstrated that this monoclonal antibody could specifically bind to hCGβ. It was also demonstrated that 6H1 monoclonal antibody could recognize the hCGβ molecules expressed on various tumor cell lines of different origins.