论文部分内容阅读
目的: 构建并表达抗人CD3嵌合抗体, 以克服鼠源单克隆抗体(mAb)用于临床的局限性。方法: 采用基因工程技术, 将抗体VL、VH 基因分别克隆入嵌合抗体表达载体VLExpress及VHExpress中。共转染COS- 7细胞后, 用ELISA检测培养上清中嵌合抗体的表达水平; 采用ProteinA亲和层析法纯化抗体, 并进行Westernblot鉴定。用FACS检测嵌合抗体结合抗原的活性; 混合淋巴细胞培养检测抗体的功能。结果: 成功地构建了VH Express VH 及VLExpress VL表达载体并表达纯化。Westernblot的结果显示, 表达的抗人CD3抗体为人鼠嵌合抗体。FACS的结果显示, 该抗体具有良好的结合抗原活性; 混合淋巴细胞培养结果显示, 该抗体具有一定的免疫抑制功能。结论: 成功地构建、表达了抗人CD3嵌合抗体, 为进一步的研究打下了基础。
OBJECTIVE: To construct and express anti-human CD3 chimeric antibody to overcome the clinical limitations of murine monoclonal antibody (mAb). METHODS: The VL and VH genes of the antibodies were cloned into the chimeric antibody expression vectors VLExpress and VHExpress, respectively, using genetic engineering techniques. After co-transfection of COS-7 cells, the expression of chimeric antibody in the culture supernatant was detected by ELISA. The antibodies were purified by Protein A affinity chromatography and identified by Western blot. The activity of the chimeric antibody binding antigen was detected by FACS; the function of the antibody was tested by mixed lymphocyte culture. Results: The VH Express VH and VLExpress VL expression vectors were successfully constructed and purified. The result of Western blot shows that the expressed anti-human CD3 antibody is a human mouse chimeric antibody. FACS results showed that the antibody has a good binding activity of antigen; mixed lymphocyte culture results showed that the antibody has a certain immunosuppressive function. Conclusion: The successful construction and expression of anti-human CD3 chimeric antibody laid the foundation for further research.