鼠抗人OX40功能性单克隆抗体的研制及其生物学特性的鉴定  

Preparation of anti-human OX40 functional monoclonal antibody and study of its biological characteristics

作　　者：孙建军[1] 王勤[1] 陈永井[1] 居颂文[1] 邱玉华[1] 施勤[1] 于葛华[1] 时文彪[1] 张学光[1] 

机构地区：[1]苏州大学生物技术研究所江苏省临床免疫重点实验室,苏州215007

出　　处：《中国免疫学杂志》2005年第6期403-405,410,共4页Chinese Journal of Immunology

基　　金：本课题为国家973重大基础研究项目(2001CB51003)国家自然科学基金重点项目(300330540)

摘　　要：目的:研制功能性鼠抗人OX40单克隆抗体。方法:以转人OX40的转基因细胞L929-OX40为免疫原,常规免疫6-8周龄的雌性BALB/c小鼠;采用B淋巴细胞融合技术,将免疫小鼠脾脏细胞与SP2/0融合,以L929-OX40转基因细胞及PHA活化的T细胞为抗体筛选阳性细胞,经免疫荧光标记分析对杂交瘤进行反复筛选和多次的克隆化培养;采用快速定性试纸法及竞争抑制结合试验分析了该单抗的亚类及抗原识别位点;采用MTT法分析单抗在体外对T细胞的促增殖效应以及ELISA分析活化T细胞分泌的细胞因子。结果:获得1株持续、稳定分泌鼠抗人OX40单克隆抗体的杂交瘤细胞株(命名为7E11),该单抗能特异性地识别人OX40分子和介导有效的共刺激信号,体外促进活化的T细胞增殖和细胞因子的分泌。结论:成功研制成一株能分泌功能性鼠抗人OX40单克隆抗体的杂交瘤,该抗体特异性地识别人OX40分子并具有在体外协同刺激T细胞的作用。Objective:To prepare a functional mouse anti-human OX40 monoclonal antibody. Methods:6-8 w female BALB/c mice were immunized with OX40 transfectant(L929-OX40) as immunogen. Mice spleen B cells were fused with SP2/0. Hybridoma cells were screened with L929-OX40 transfectant and PHA activated T cells by FCM. Fast-strip analysis was performed to identify Ig subclass of this mAb. The epitope recognized by this mAb was studied by OX40-FTTC(ACT35) competitive assay. Proliferation and cytokine secretion of T cells in vitro were studied by MTT assay and EI.TSA respectively.Results:One hybridoma cell line named 7E11 was obtained, which shows a property of secreting anti-human OX40 monoclonal antibody(mAb) continuously and steadily.The results show this mAb can specifically recognize human 0X40 molecule and mediate effective costimulatory signal.It can promote the proliferation of activated T cells and cytokine secretion in vitro. Conclusion:One hybridoma cell line which can secret functional mouse anti-human OX40 mAb was developed successfully.This mAb can recognize human OX40 specifically and conduct costimulatory effect for T cells in vitro.

关 键 词：OX40 杂交瘤 单克隆抗体 T细胞 

分 类 号：R392.12[医药卫生—免疫学]