抗沙眼衣原体MIP蛋白单克隆抗体的制备及特性鉴定  

Production and identification of monoclonal antibodies against MIP protein of Chlamydia trachomatis

作　　者：陆春雪[1] 彭波[2] 陈超群[1] 周洲[1] 彭江南[1] 钟光明[1] 吴移谋[1] 

机构地区：[1]南华大学医学院病原生物学研究所,衡阳421001 [2]南华大学医学院肿瘤研究所,衡阳421001

出　　处：《中国免疫学杂志》2014年第1期80-84,共5页Chinese Journal of Immunology

基　　金：国家自然科学基金(No.81202374);湖南省自然科学基金(No.12JJ3113);湖南省高校科技创新团队(湘教通[2010]212号)等基金资助

摘　　要：目的:制备小鼠抗沙眼衣原体MIP蛋白单克隆抗体并鉴定其生物学特性。方法:重组MIP蛋白免疫BALB/c小鼠,取小鼠脾细胞与SP2/0骨髓瘤细胞进行常规融合,间接ELISA法筛选阳性克隆并进行有限稀释克隆化,建立分泌抗MIP蛋白单克隆抗体的杂交瘤细胞株,进一步采用ELISA法鉴定单克隆抗体的效价、类别及抗MIP蛋白特异性,IFA法鉴定单克隆抗体的衣原体种属特异性及中和能力。结果:建立了2株能稳定分泌抗MIP蛋白单克隆抗体的杂交瘤细胞株(1R6H6、2L9D2),ELISA法测得培养上清效价均为1:1 600,免疫球蛋白类型分别为IgG2a和IgG1;2株单克隆抗体不仅能特异性的识别MIP重组蛋白,而且能特异性识别沙眼衣原体血清型A、D、L2所编码的内源性MIP蛋白,但不识别其他衣原体优势蛋白及MoPn、AR39和6BC;体外中和试验表明,2株单克隆抗体均能有效中和衣原体的感染性。结论:获得了有活性的能特异性识别MIP蛋白的单克隆抗体,为深入研究MIP蛋白的结构和功能奠定了基础。Objective ：To produce monoclonal antibodies （mAbs） against MIP protein of Chlamydia trachomatis and prelimina- ry analysis of their biological characteristics. Methods： BALB/c mice were regularly immunized with purified MIP protein and the spleen cells were collected and fused with SP2/0 mouse myeloma cells. The positive hybridoma cell lines were screened by ELISA and subeloned 3 times by limiting dilution culture. ELISA was further carried out to determine the mAbs titers, isotypes, as well as the spe- cificities to MIP protein. Indirect immunofluorescence assay （IFA） was performed to identify the chlamydial species specificity and neutralization activity. Results： Two hybridoma cell lines were obtained and named （1R6H6,2L9D2） , ELISA showed that the titers of these mAbs were 1：1 600, and the isotypes were belonged to IgG2a and IgG1. Both mAbs reacted strongly with the purified GST-MIP protein, as well as endogenous MIP protein expressed by Chlamydia trachomatis serovar A, D and I2, but not other chlamydial immunodominant proteins and Chlamydia muridarum （MoPn） , Chlamydia pneumonia AR39, Chlamydia psittaci 6BC. The neutralization test showed that these mAbs can neutralize chlamydial infections in vitro. Conclusion： 2 mAbs against MIP protein have been successfully produced with high specificity and activity, which will certainly facilitate further study on MIP protein structure and function.

关 键 词：沙眼衣原体 MIP蛋白 单克隆抗体 鉴定 

分 类 号：R392.7[医药卫生—免疫学]