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为了表达具有中和活性的抗禽流感H5N1病毒人-鼠嵌合IgA抗体,采用RT-PCR法克隆具有中和活性的抗禽流感H5N1-HA鼠源单克隆抗体的轻重链可变区基因及相应的信号肽编码序列,分别与人免疫球蛋白IgA2重链恒定区、Kappa恒定区基因拼接,构建表达质粒pEF-IGHA9和pEF-IGK9,共转染二氢叶酸还原酶缺陷型CHO(CHO-dhfr-)细胞,用ELISA检测培养上清中嵌合IgA抗体的表达,对纯化的嵌合抗体进行SDS-PAGE、Western blotting印迹分析。结果成功地在CHO细胞中表达了抗禽流感H5N1病毒人-鼠嵌合IgA抗体,为制备抗H5N1重组分泌型IgA预防性抗体制剂奠定了良好的基础。
To express neutralizing human anti-H5N1 human chimeric IgA antibody against avian influenza virus, RT-PCR was used to clone the light chain heavy chain variable region genes of mouse monoclonal antibody against H5N1-HA with neutralizing activity and The corresponding signal peptide coding sequences were spliced with the heavy chain constant region of human immunoglobulin IgA2 and Kappa constant region respectively to construct expression plasmids pEF-IGHA9 and pEF-IGK9, co-transfected with dihydrofolate reductase-deficient CHO (CHO- dhfr-) cells. The expression of chimeric IgA antibody in the culture supernatant was detected by ELISA. The purified chimeric antibody was analyzed by SDS-PAGE and Western blotting. Results The human anti-H5N1 virus human-mouse chimeric IgA antibody was successfully expressed in CHO cells, which laid a good foundation for the preparation of anti-H5N1 recombinant secretory IgA antibody.