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目的建立能够分泌特异性抗小鼠膜联蛋白A2(AnxA2)的大鼠-小鼠融合单克隆抗体杂交瘤细胞系。方法利用重组小鼠AnxA2蛋白作为抗原免疫Wistar大鼠,通过细胞融合和杂交瘤筛选技术建立特异分泌抗小鼠AnxA2蛋白的大鼠-小鼠融合单抗杂交瘤细胞株。采用酶联免疫吸附方法确定单克隆抗体亚型,免疫印迹和流式细胞术验证所筛选的单克隆抗体的特异性。结果通过对30多个分泌抗体的融合细胞克隆的筛选,获得一株分泌特异性抗小鼠AnxA2的大鼠-小鼠融合单克隆抗体杂交瘤细胞系SZ-158,连续培养传代后染色体分析证实,Wistar大鼠脾细胞与小鼠SP2/0细胞稳定融合;免疫印迹和流式细胞术显示该单克隆抗体可以特异识别小鼠AnxA2重组蛋白。结论该研究所获得的大鼠-小鼠融合单克隆抗体特异性针对小鼠AnxA2蛋白,可用于AnxA2转基因小鼠中AnxA2表达的检测,为深入研究AnxA2的功能提供了物质保障。
OBJECTIVE To establish a rat-mouse fusion monoclonal antibody hybridoma cell line capable of secreting a specific anti-mouse annexin A2 (AnxA2). Methods Wistar rats were immunized with recombinant mouse AnxA2 protein. The hybridoma cell lines secreting anti-mouse AnxA2 protein were established by cell fusion and hybridoma screening. Monoclonal antibody subtypes were determined by enzyme-linked immunosorbent assay, and the specificity of the selected monoclonal antibodies was verified by Western blotting and flow cytometry. Results A rat-mouse fusion monoclonal antibody-secreting hybridoma cell line SZ-158 secreting anti-mouse AnxA2 was obtained through screening of more than 30 antibody-secreting fusion cell clones. Chromosome analysis after continuous culture confirmed The splenocytes of Wistar rats were stably fused with mouse SP2 / 0 cells. Western blotting and flow cytometry showed that the monoclonal antibodies could specifically recognize mouse AnxA2 recombinant protein. Conclusion The rat-mouse fusion monoclonal antibody obtained from this study is specific for mouse AnxA2 protein and can be used for the detection of AnxA2 expression in AnxA2 transgenic mice, which provides a material guarantee for further study on the function of AnxA2.