title: [Expression of human Fab antibody against keratin in E.coli and its renaturation].
abstract: OBJECTIVE
  To express Fd fragment and L chain of human anti-keratin Fab in E.coli BL21(DE3) respectively and obtain human anti-keratin Fab by renaturation in-vitro.
  
  
  METHODS
  Genes of L chain and Fd fragment of anti-keratin antibody from the plasmids p3MH/ Fab were subcloned into vector pET32a respectively. The recombinant plasmids pETL and pETFd were transformed into E.coli BL21(DE3) and induced to express with IPTG. Fd fragment and L chain inclusion bodies were solubilized and combined in equal molar ratio in the refolding solution. The renatured Fab was characterized by SDS-PAGE, Western blot and ELISA.
  
  
  RESULTS
  Fd fragment and L chain of human anti-keratin Fab were efficiently expressed. ELISA and Western blot showed that the renatured Fab could bind with human keratin.
  
  
  CONCLUSIONS
  The successfully prepared anti-keratin Fab with binding activity to human keratin laid a solid foundation for its further application.
authors.affiliation: Dermatology Center of Chinese PLA, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China. fanjianyongfmmu@sina.com
authors.forename: Jian-yong
authors.initials: JY
authors.lastname: Fan
authors.forename: Gang
authors.initials: G
authors.lastname: Wang
authors.forename: Wei
authors.initials: W
authors.lastname: Li
authors.forename: Yuan-ming
authors.initials: YM
authors.lastname: Wu
authors.forename: Yu-feng
authors.initials: YF
authors.lastname: Liu
mesh_terms: D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000235:genetics / Q000378:metabolism*; D007128:Immunoglobulin Fragments / Q000096:biosynthesis* / Q000235:genetics; D007147:Immunoglobulin Light Chains / Q000096:biosynthesis* / Q000235:genetics; D002479:Inclusion Bodies / Q000378:metabolism; D007633:Keratins / Q000235:genetics / Q000276:immunology*; D020673:Protein Renaturation; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D014170:Transformation, Genetic
chemical_list: D007140:Immunoglobulin Fab Fragments; D007128:Immunoglobulin Fragments; D007147:Immunoglobulin Light Chains; D011993:Recombinant Fusion Proteins; C100277:immunoglobulins, Fd; D007633:Keratins