抗人CD28鼠-人嵌合抗体的构建及瞬时表达

Construction and transient expression of the mouse-human chimeric antibody against human CD28

目的构建及瞬时表达抗人CD28鼠-人嵌合抗体。方法采用基因工程技术,从分泌鼠抗人CD28单克隆抗体的杂交瘤细胞株(克隆号:2F5)中克隆出抗体重链和轻链可变区基因及相应信号肽编码序列。分别与人免疫球蛋白IgG1恒定区重、轻链基因拼接,构建含有重组基因的pIRES表达质粒pIRES/h2F5。转染293T细胞并用抗性药物G418选择培养。采用流式细胞术,检测瞬时转染细胞后嵌合抗体的表达情况。结果抗人CD28鼠人嵌合抗体得到表达并能与CD28分子特异性结合。结论抗人CD28鼠-人嵌合抗体的构建和瞬时表达是成功的,为后续构建稳定分泌抗人CD28鼠-人嵌合抗体的CHO基因转染细胞株奠定了基础。

Purpose To construct and transiently express of a mouse-human chimeric antibody against human CD28 molecule. Methods By using genetic technology, total RNA was extracted from the murine hybridoma cell line （Clone number： 2F5） secreting anti-CD28 monoclonal antibody. The variable region and the correspond signal peptide coding sequence were cloned, then fused with the coding sequence of the constant region of the human IgG1 heavy chain and light chain respectively. The mouse-human chimeric antibody recombinant plasmid was established. 293T cell line was transfected with the recombinant plasmid and the transfectants were selected by G418 contained medium. Using FACS to detect the chimeric antibody＇s transient expression level. Results The chimeric antibody against human CD28 was expressed and recognized the CD28 molecular specifically. Conclusions Chimeric antibody is successfully constructed and is transiently expressed, thus a foundation is laid for establishing CHO cell line secreting chimeric anti-CD28 monoclonal antibody.