雪卡毒素单克隆抗体的制备及鉴定  

Preparation and identification of monoclonal antibodies against ciguatoxin

作　　者：张彩霞[1] 闫鸿鹏[1] 郑杰[1] 周丽华[1] 张焜[1] 赵肃清[1] 

机构地区：[1]广东工业大学轻工化工学院

出　　处：《免疫学杂志》2011年第10期890-893,共4页Immunological Journal

基　　金：国家自然科学基金(20872020;20672023);广东省自然科学基金(8251009001000005)

摘　　要：目的制备雪卡毒素的单克隆抗体并对其生物学特性进行鉴定。方法以雪卡毒素类似物莫能菌素为半抗原,分别采用混合酸酐法将其与载体牛血清白蛋白(BSA)偶联合成免疫抗原M-BSA,碳二亚胺法与卵清白蛋白(OVA)偶联合成包被抗原M-OVA。以M-BSA免疫Balb/c小鼠,取其脾细胞与Sp2/0骨髓瘤细胞融合制备杂交瘤。以自制雪卡毒素提取物为竞争抗原,间接竞争ELISA法筛选稳定分泌雪卡毒素抗体的特异性细胞株。小鼠体内诱生腹水,辛酸硫酸铵沉淀法进行纯化。采用抗体亚型鉴定试剂盒鉴定所得抗体的亚型,间接竞争ELISA法鉴定抗体的特异性和交叉性。结果获得3株稳定分泌抗雪卡毒素抗体的杂交瘤细胞株1D5、2G11、3G11,其Ig亚型均属于IgG1亚型。交叉反应结果表明3株单克隆细胞产生的抗体除与莫能菌素有较高的交叉反应外,与其他雪卡毒素类似物均无明显交叉反应。结论成功筛选到了分泌雪卡毒素抗体的细胞株,为雪卡毒素免疫分析方法的进一步研究奠定基础。This study aims to prepare monoclonal antibodies（McAbs） against ciguatoxin and to characterize their properties.Mixed anhydride and EDC methods were used for coupling monensin with BSA/OVA to prepare immunogen and coating antigen of CTXs respectively.Balb/c mice were immunized with artificial ciguatoxin antigen M-BSA,and hybridoma was generated with immunized mouse spleen cells and Sp2/0 myeloma cells.The hybridoma cell lines which secret McAbs against ciguatoxin were selected and subcloned 4 times.Consequently,3 hybridoma cell lines secreting McAbs against ciguatoxin were obtained and named as 1D5,2G11 and 3G11.Then the McAbs were purified from the mouse ascites followed by characterization with enzyme-linked immunosorbent assay（ELISA）.All the three McAbs were belonged to IgG1 subtype.McAbs against ciguatoxin can bind with ciguatoxin specifically,thus which can be useful for sensitive detection of ciguatoxin.

关 键 词：雪卡毒素 莫能菌素 单克隆抗体 间接竞争ELISA 

分 类 号：R392[医药卫生—免疫学]