title: [Expression and immunological activity of an anti-CD3×VEGFR2 bispecific single-chain antibody].
abstract: OBJECTIVE
  To construct and express an anti-VEGFR2/anti-CD3 bispecific single-chain antibody (bscVEGFR2×CD3)and to identify its binding specificities to CD3 and VEGFR2.
  
  
  METHODS
  The gene encoding anti-VEGFR2/anti-CD3 bispecific single-chain antibody was designed and synthesized. Bispecific single-chain antibody (bsc-Ab) DNA was subcloned into a eukaryotic expression vector pcDNA3.1(+), then transfected into Chinese hamster ovary (CHO) cells and stable expression cell lines were selected. Expressed Bsc-Ab was purified by His-tag affinity chromatography and confirmed by 120 g/L SDS-PAGE and Western blotting. Antigen binding activity of the bsc-Ab was analyzed by FACS.
  
  
  RESULTS
  The plasmid DNA containing bispecific single-chain fragments were confirmed. BscVEGFR2×CD3 was secreted by CHO into the supernatant. Six stable expression cell lines were established. The molecular weight of bsc-Ab was correct indicated by SDS-PAGE and Western blotting. The bsc-Ab could specifically bind to CD3(+); jurkat cells and VEGFR2(+); A375 cells.
  
  
  CONCLUSIONS
  An anti-VEGFR2/anti-CD3 bispecific single-chain antibody is successfully constructed and expressed, and the antibody has specific binding capacity to CD3 and VEGFR2.
authors.affiliation: Department of Pharmacy, College of Pharmacy, Harbin Medical University, Harbin 150081, China.
authors.forename: Li-juan
authors.initials: LJ
authors.lastname: Dai
authors.forename: Cheng-wei
authors.initials: CW
authors.lastname: Yan
authors.forename: Shu-zhen
authors.initials: SZ
authors.lastname: Li
authors.forename: Li
authors.initials: L
authors.lastname: Chen
authors.forename: Xin-yu
authors.initials: XY
authors.lastname: Li
authors.forename: Rui-hui
authors.initials: RH
authors.lastname: Shan
authors.forename: Xue-wei
authors.initials: XW
authors.lastname: Yu
authors.forename: Xiao-bo
authors.initials: XB
authors.lastname: Tang
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D017252:CD3 Complex / Q000276:immunology*; D016466:CHO Cells; D045744:Cell Line, Tumor; D006224:Cricetinae; D003412:Cricetulus; D005786:Gene Expression Regulation / Q000276:immunology*; D023061:Gene Order; D006801:Humans; D019169:Jurkat Cells; D020014:K562 Cells; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000037:antagonists & inhibitors / Q000276:immunology*
chemical_list: D018033:Antibodies, Bispecific; D017252:CD3 Complex; D011993:Recombinant Fusion Proteins; D040301:Vascular Endothelial Growth Factor Receptor-2