抗人T淋巴细胞CD4人-鼠嵌合抗体的真核表达  

Eukaryotic Expression of Human/Mouse Chimeric Antibody against Human T Lymphocyte CD4

作　　者：张雪[1] 张爱华[1] 闭兰[1] 赵亚杰[1] 孙可芳[1] 方志正[1] 

机构地区：[1]武汉生物制品研究所免疫学研究室,武汉430060

出　　处：《中国生物制品学杂志》2009年第10期945-948,952,共5页Chinese Journal of Biologicals

摘　　要：目的在小鼠骨髓瘤细胞SP2/0中表达抗人T淋巴细胞CD4人-鼠嵌合抗体。方法真核表达质粒PAG4622+CD4VL和PAH4604+CD4VH经酶切和序列测定后,采用电穿孔转染技术将二者共转染SP2/0细胞,经组胺醇和霉酚酸联合筛选阳性克隆,通过ELISA、流式细胞术、RT-PCR和DNA测序的方法进行初步鉴定。结果真核表达质粒经酶切和测序鉴定证明构建正确。获得2株分泌抗人T淋巴细胞CD4人-鼠嵌合抗体的阳性SP2/0细胞克隆0925CASP2/0和1107CASP2/0,嵌合抗体表达量为0.5～2ng/ml。结论已成功地在SP2/0细胞中表达了抗人T淋巴细胞CD4人-鼠嵌合抗体,为该抗体在其他真核细胞中的高效表达及临床应用奠定了基础。Objective To express human/mouse chimeric antibody against human T lymphocyte CD4 in murine myeloma SP2/0 cells.Methods Eukaryotic expression vectors PAG4622 ＋ CD4VL and PAH4604 ＋ CD4VH were identified by restriction analysis and sequencing,then co-transfected to SP2/0 cells by electroporation.The positive clones were screened by histidinol and mycophenolic acid and preliminarily identified by ELISA,flow cytometry,RT-PCR and DNA sequencing.Results Both restriction analysis and sequencing proved that eukaryotic expression vectors PAG4622 ＋ CD4VL and PAH4604 ＋ CD4VH were constructed correctly.Two SP2 /0 cell clones secreting human/mouse chimeric antibody against human T lymphocyte CD4,i.e.0925CASP2 /0 and 1107CASP2/0,were screened,in which the expression level of chimeric antibody was 0.5 ～ 2 ng /ml.Conclusion Human /mouse chimeric antibody against human T lymphocyte CD4 was successfully expressed in murine myeloma SP2 /0 cells,which laid a foundation of high expression in other eukaryocytes and clinical application of the chimeric antibody.

关 键 词：CD4 嵌合抗体 电穿孔 SP2/0细胞 真核表达 

分 类 号：Q786[生物学—分子生物学]