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目的 制备抗GPA单克隆抗体以建立“GPA体细胞突变分析法”。方法 用O ,N型人红细胞 ,以及纯化人GPAN 免疫BALB/c小鼠 ,经杂交瘤技术制备 ,用细胞凝集、间接免疫荧光筛选 ,用ELISA、Westernblot、酶处理红细胞的间接免疫荧光标记鉴定。结果 获得抗GPAN 单克隆抗体 6A8杂交瘤细胞系。其单抗属lgG3亚类 ,λ型轻链 ,识别和结合GPAN 氨基端 1～ 8位决定位点 ,对糖链依赖。流式细胞分选结果显示 6A8特异结合二甲亚胺固定的人MN和N型红细胞。结论 对分析由于基因突变和糖基化变异所致N ,MN红细胞GPAN 突变有理论研究和应用价值。
Objective To prepare anti-GPA monoclonal antibody to establish “GPA somatic mutation assay”. Methods BALB / c mice were immunized with O, N type human erythrocytes and purified human GPAN. The hybridomas were prepared by hybridoma technique. Indirect immunofluorescent labeling was used to identify the erythrocytes by ELISA, Western blot and ELISA. As a result, an anti-GPAN monoclonal antibody 6A8 hybridoma cell line was obtained. Its monoclonal antibody lgG3 subclass, λ light chain, recognition and binding GPAN amino terminal 1 to 8 decision sites, the sugar chain. Flow cytometry results show that 6A8 specifically binds to dimethylidine-immobilized human MN and N-type erythrocytes. Conclusion There are theoretical and practical value in the analysis of GPAN mutation of N, MN erythrocytes due to gene mutation and glycosylation mutation.