Stable expression of ICOS/Fc in CHO cells

    DU Junª²Feng1, WANG Weiª²Zhong1, ZHUANG Ran2, WANG Chunª²Yan2, GUAN Wenª²Xian1

    1Department of Gastrointestinal Surgery, Xijing Hospital, 2Deª²partment of Immunology, School of Basic Medicine, Fourth Military Medical University, Xi¬ðan 710033, China

    ¡¾Abstract¡¿ AIM: To study the stable expression of pICOS/Fc fusion protein expression vector in Chinese hamster ovary (CHO) cells and to purify and identify its expression product in vitro. METHODS: Expression vector pICOS/Fc and resistance plasmid pEGFP were cotransfected into CHO cells with electroporation. The selective medium containing G418 was then employed to select the positive colony. After repeating cloning culture and double antibody sandwich ELISA detection, the cell clones with stable expression were obtained. After purified with protein A affinity chromatography, the product was studied with SDSª²PAGE and Western blotting to determine its molecular weight, purity and antigen specificity. RESULTS: CHO cell clones stably expressing pICOS/Fc fusion proteins were obtained and the fusion protein was available from the supernatant of cell culture media after purified with protein A affinity chromatography. A protein band with molecular weight of 70 KDa was found in the SDSª²PAGE. This protein specifically bound with antiª²human Fc monoclonal antibody labeled HRP. CONCLUSION: Recombined mouse ICOS/Fc fusion protein with antigen specificity is successfully and stably expressed in CHO cells. ......