title: [Construction and expression of anti-human integrin alphavbeta3 scFv].
abstract: OBJECTIVE
  To construct single chain antibody (scFv) gene of mAb E10 against human integrin alphavbeta3.
  
  
  METHODS
  The VH and VL genes were amplified from hybridoma cells secreting mAb E10 by RT-PCR and connected with the use of linker (Gly4Ser)3 to assemble scFv gene. The scFv gene was cloned into prokaryotic expression vector pTIG-TRX and expressed in E. coli BL21 (DE3).
  
  
  RESULTS
  SDS-PAGE analysis showed the expressed recombinant protein with relative molecular mass (Mr) being 31,000. Western blot confirmed that the protein was labeled with His6. scFv protein was expressed as soluble protein under the condition of a small amount of IPTG induction and culture at lower temperature. The purity of the protein purified through Ni-NTA agarose metal affinity resin column was over 91%. The purified protein could bind to the human integrin alphavbeta3 by ELISA confirmation.
  
  
  CONCLUSIONS
  scFv against human integrin alphavbeta3 has been successfully constructed and expressed,which lays the foundation for further clinical research.
authors.affiliation: Institute of Microbiology & Epidemiology, Academy of Military Medical Sciences, Beijing 100071, China. wangchen2001@163.com
authors.forename: Chen
authors.initials: C
authors.lastname: Wang
authors.forename: Li-hua
authors.initials: LH
authors.lastname: Hou
authors.forename: Yan-ming
authors.initials: YM
authors.lastname: Zhang
authors.forename: Jian-min
authors.initials: JM
authors.lastname: Li
authors.forename: Zhen-lin
authors.initials: ZL
authors.lastname: Liao
authors.forename: Gui-xin
authors.initials: GX
authors.lastname: Du
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.forename: Qi-hong
authors.initials: QH
authors.lastname: Sun
authors.forename: Yi-gang
authors.initials: YG
authors.lastname: Tong
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000378:metabolism; D050438:Genes, Immunoglobulin Heavy Chain; D050439:Genes, Immunoglobulin Light Chain; D005822:Genetic Vectors; D006801:Humans; D006825:Hybridomas / Q000378:metabolism; D007128:Immunoglobulin Fragments / Q000096:biosynthesis* / Q000235:genetics; D039302:Integrin alphaVbeta3 / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D007128:Immunoglobulin Fragments; D039302:Integrin alphaVbeta3; D011994:Recombinant Proteins; C055966:immunoglobulin Fv