【摘要】 制备抗双链DNA(dsDNA)单克隆抗体并对其生物学特性进行初步研究。以活化淋巴细胞的DNA为免疫原,常规免疫BALB/c小鼠,采用B淋巴细胞杂交瘤技术制备单克隆抗体;用体内诱生法制备腹水,以间接酶链反应吸附试验(ELISA)法分析抗体的亚类、特异性和亲和力;用免疫荧光法检测抗体的荧光核型;用考马斯亮蓝法检测注入杂交瘤细胞的小鼠的尿蛋白含量。结果显示,成功获得了1株持续稳定分泌抗dsDNA单克隆抗体的杂交瘤细胞株,命名为6C2;该细胞株所分泌的抗体亚类为小鼠IgG2a,能特异性结合dsDNA,亲和力常数为2.67×10~8mol/L,免疫荧光核型为均质型;将6C2细胞注入正常BALB/c小鼠腹腔后,小鼠的尿蛋白含量较对照组明显增高。结果表明,抗dsDNA单克隆抗体6C2是致病性抗体,对于进一步研究抗dsDNA抗体的致病机制具有重要的价值。更多还原

【Abstract】 A functional monoclonal antibody against double-strand DNA(dsDNA)was prepared and characterized by immu- nization of BALB/c mice with ConA activated lymphocyte-derived DNA(ALD-DNA)as immunogen.By means of the standard hybridoma technique for B lymphocytes to cell fusion,one hybridoma cell line was established and designed as 6C2 clone.This clone was propagated by injection into the sensitized mice,the aseitic fluid of mice was collected and purified by affinity chro- matography with protein-G.ELISA assay was performed to identify isotype,specificity and affinity of the mAb.The pattern of immunofluorescence was performed on Hep-2 cell and the urine protein of the bybridoma-injecting mice were measured with the coomassie brilliant blue assay.The results showed this mAb was of IgG2a isotype and could specifically recognize the ds- DNA.The analysis of affinity indicated that 6C2 was of high affinity.After injecting hybridoma cells 6C2 into the sensitized mice,the urine proteins were dramatically higher than control groups.It indicates 6C2 is a pathological antibody and would be of significant value in the basic studies for the roles of anti-dsDNA antibodies in the pathogenesis of SLE.更多还原