title: [Expression, renaturation and activity of BAC5-scFv expressed as inclusion body in E.coli].
abstract: OBJECTIVE
  To study the renaturation, purification and binding activity of scFv of anti-nasopharyngeal carcinoma monoclonal antibody(mAb) BAC(5) expressed as inclusion body in E.coli.
  
  
  METHODS
  The E.coli BL21(DE3) transformed with the pET 22b-scFv was cultured and pulvereged by ultrasonic cell disintegrator. The collected inclusion bodies were denatured with 8 mol/L urea and renatured by dilution refolding, step dialysis and gel filtration chromatography. Binding activity of renatured BAC(5)-scFv was determined by immunohistochemical staining and Western blot.
  
  
  RESULTS
  BAC(5)-scFv purified though Ni-NTA His Bind chromatographic clomn showed high purity. The highest proteins recovery rate was obtained through gel filtration chromatography. It was proved by Western blot and immunocytochemical staining that the renatured BAC(5)-scFv protein could specifically bind to CNE2 cells.
  
  
  CONCLUSIONS
  BAC(5)-scFv expressed as inclusion body retained good activity after being dissolved, purified and renatured, which paves the way for preparing large amount of BAC(5)-scFv to be used for the study of radioimmunoimaging and therapy of nasopharyngeal carcinoma.
authors.affiliation: Department of Nuclear Medicine, Zhongshan Medical College, Sun yat-sen University, Guangzhou 510080, China. tcaihua2004@126.com
authors.forename: Cai-hua
authors.initials: CH
authors.lastname: Tang
authors.forename: Xi-bin
authors.initials: XB
authors.lastname: Xiao
authors.forename: Tao
authors.initials: T
authors.lastname: Wu
authors.forename: Chang-sheng
authors.initials: CS
authors.lastname: Liang
authors.forename: Chang-zheng
authors.initials: CZ
authors.lastname: Liu
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007128:Immunoglobulin Fragments / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007150:Immunohistochemistry; D002479:Inclusion Bodies / Q000378:metabolism*; D009303:Nasopharyngeal Neoplasms / Q000276:immunology; D017510:Protein Folding
chemical_list: D000911:Antibodies, Monoclonal; D007128:Immunoglobulin Fragments