抗人DcR3单克隆抗体的制备、鉴定及应用  

Preparation,characterization,and application of monoclonal antibodies against human decoy receptor 3

作　　者：刘瑞振[1] 张长弓[1] 何琼[2] 苏金华[1] 陈彩霞[1] 陈福[1] 庄国洪[1] 

机构地区：[1]厦门大学医学院抗癌研究中心 [2]厦门市波生生物技术有限公司

出　　处：《免疫学杂志》2009年第1期34-38,共5页Immunological Journal

基　　金：厦门大学科研启动资金(Z03103);厦门市科技计划项目(3502Z20083008)资助

摘　　要：目的制备抗DcR3单克隆抗体(mAb),鉴定其生物学特性,并应用于ELISA、Western blot、Flowcytometry(FCM)检测。方法以纯化的可溶性DcR3(sDcR3)免疫Balb/c小鼠,采用杂交瘤技术制备抗DcR3 mAb。用Ig亚类ELISA试剂盒鉴定抗DcR3mAb的亚类。用ELISA方法测定抗DcR3 mAb与sDcR3结合的特性,SDS-PAGE鉴定抗DcR3 mAb与SW480细胞上清中DcR3结合的特性,以鉴定mAb的特性。用间接ELISA法检测腹水mAb的效价、亲和常数并进行表位分析。Western blot检测mAb的特异性及应用,并用所获抗DcR3单克隆抗体(mAb)通过流式细胞仪检测肿瘤细胞表面DcR3的表达水平。结果获得4株可分泌DcR3 mAb的杂交瘤细胞系ZZ-393、ZZ-394、ZZ-151和ZZ-268。其中DcR3 mAb ZZ-268(下文简称为ZZ-268)的Ig亚类为IgG1(κ型);腹水效价为1×10-5;亲和常数为1.28×109水平;ZZ-268和ZZ-151可识别与其他2种抗体不同的抗原表位;Western blot证实,获得的ZZ-268可特异性地识别DcR3;通过流式细胞术可敏感地检测到不同肿瘤细胞表面DcR3的表达水平。结论获得4株抗DcR3的mAb,其中ZZ-268效价高、特异性强,将此抗体用于膜DcR3与sDcR3的检测分析。Objective To prepare monoclonal antibodies mAb against DcR3,characterize their biological properties and apply them to ELISA,Flow cytometry,and Western blot detection.Methods Balb/c mice were immunized with soluble DcR3(sDcR3) for preparing mAb by using hybridoma technique.Ig subclass and specificity of mAbs were analyzed by ELISA and SDS-PAGE,respectively.The titres,relative affinity,and epitopes of mAbs in ascitic fluid were determined by indirect ELISA.The specificity and application of mAb were detected...

关 键 词：DCR3 单克隆抗体 制备 

分 类 号：R392.11[医药卫生—免疫学]