[Expression, identification and bioactivity characterization of an anti-CD3/anti-CD20 bispecific single-chain antibody].
The synthetic gene with 1640bp encoding for the anti-CD3/anti-CD20 bispecific single-chain antibody was designed and obtained by SOE (splicing by overlap extension) PCR. The cDNA was cloned into Flp-In expression vector pcDNA5/FRT and transfeced into Flp-In CHO cells to generate a stable expression cell line with a capacity for expressing anti-CD3/anti-CD20 bispecific single-chain antibody at 300 microg/L. The protein, which had a molecular weight of about 70 kD,was purified by Ni-NTA affinity chromatography and identified by SDS-PAGE and Western-blot analysis. Immunofluorescence assay and cellular rosetting showed that it can react specifically on Jurkat (CD3+) and Ramous (CD20+) cells. The lysis of human PBL against CD20-positive lymphoma Ramous cells in the presence of the anti-CD3/anti-CD20 bispecific single-chain antibody can observed by microscope. All these results would lighten the further study of its biological functions in vitro and in vivo.
Yu
Rui
R
Institute of Biotechnology, Academy of Military Medical Sciences, Beijing 100071, China.
Li
Shi-Chong
SC
Wu
Ben-Chuan
BC
Liu
Hong
H
Ye
Ling-Ling
LL
Chen
Zhao-Lie
ZL
D000818:Animals; D018033:Antibodies, Bispecific / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology; D018951:Antigens, CD20 / Q000276:immunology*; D017252:CD3 Complex / Q000276:immunology*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D016393:Lymphoma, B-Cell / Q000378:metabolism; D011994:Recombinant Proteins; D014162:Transfection; D014407:Tumor Cells, Cultured
D018033:Antibodies, Bispecific; D000911:Antibodies, Monoclonal; D018951:Antigens, CD20; D017252:CD3 Complex; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins