一株识别CD40分子新位点的单克隆抗体研制及生物学特性鉴定  

Preparation of a novel functional anti-human CD40 monoclonal antibody and identification of its bioactivity

作　　者：汤琳[1] 郑璐[1] 戚春建[1] 马泓冰[1] 周璇[1] 庄羽美[1] 罗先富[1] 徐颖[1] 张学光[1] 

机构地区：[1]苏州大学医学生物技术研究所江苏省医学临床免疫学重点实验室,江苏苏州215007

出　　处：《细胞与分子免疫学杂志》2007年第9期841-843,846,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金面上项目(30571690);教育部博士点专项基金项目(20050285014)

摘　　要：目的：研制识别CD40分子新位点的单克隆抗体（mAb）。方法：以转人CD40转基因细胞L929-CD40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-CD40转基因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mAb的亚类及抗原识别位点；采用MTT法分析mAb在体外对肿瘤细胞的促增殖效应、ELISA法测定细胞因子分泌以及PI染色分析细胞周期。结果：获得1株持续、稳定分泌鼠抗人CIM0mAb的杂交瘤细胞株（命名为286），该mAb能特异性地识别人CD40分子并较好的识别肿瘤细胞株H08910表达的CD40分子并能够体外促进肿瘤细胞增殖。结论：成功地研制1株识别CD40新位点mAb，该mAb能够很好地识别肿瘤细胞表达的CD40分子并具有体外促进肿瘤细胞生长的作用。AIM： To prepare a novel functional mouse anti-human CD40 monoclonal mAb. METHODS： Female BALB/c mice of 6 - 8 weeks old were immunized with CD40 transfectant （L929-CD40） as immunogen. The spleen B cells of the mice were fused with Sp2/0. The hybridoma cells were screened with CD40 transfectant （L929-CD40） by FCM. Fast-strip analysis was performed to identify Ig subclass of this mAb. The epitope recognized by this mAb was detected by Bio/SC11 competitive assay. The proliferation and cell cycle of tumor cells in vitro were studied by MTT assay and PI staining respectively. RESULTS： One hybridoma cell line named 2B6 was obtained, which had the property of secreting anti-human CD40 monoclonal antibody continuously and steadily. This mAb specifically recognized human CD40 molecule and promoted the proliferation of tumor cells in vitro. CONCLUSION： One hybddoma cell line which can secret a novel functional mouse anti-human CD40 mAb has been developed successfully. This mAb can specifically recognize human CD40 and influence the growth of tumor cells in vitro.

关 键 词：CD40 杂交瘤 单克隆抗体 肿瘤细胞 

分 类 号：R392.12[医药卫生—免疫学]