title: [Secretory expression of chimeric Fab antibody HAb18 against human hepatocellular carcinoma in Pichia pastoris].
abstract: OBJECTIVE
  To express secretively chimeric Fab antibody HAb18 (cFab) against human hepatocellular carcinoma in Pichia pastoris.
  
  
  METHODS
  Genes encoding CL chain and Fd fragment of cFab antibody HAb18 were subcloned into vectors pPIC9K and pPICZalphaA, respectively. After confirmed by DNA sequence analysis, the recombinant plasmids pPIC9K/CL and pPICZalphaA/Fd were transformed into the genome of Pichia pastoris GS115. Mut(+) multiple insert transformants were screened by G418 and Zeocin and then induced with 5 mL/L methanol to express cFab.
  
  
  RESULTS
  4 days after methanol induction, 26 mg/L of the cFab fragment was detected in the culture supernatant. Western blot proved that the expressed protein could specifically bind with HAb18GEF antigen.
  
  
  CONCLUSIONS
  The successful expression of cFab/HAb18 in Pichia pastoris lays the foundation for large-scale production and further application of the antibody.
authors.affiliation: Department of Hepatobiliary Surgery, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
authors.forename: Hong-lin
authors.initials: HL
authors.lastname: Dong
authors.forename: Jin-liang
authors.initials: JL
authors.lastname: Xing
authors.forename: Jia-ze
authors.initials: JZ
authors.lastname: An
authors.forename: Xiang-min
authors.initials: XM
authors.lastname: Yang
authors.forename: Xi-ying
authors.initials: XY
authors.lastname: Yao
authors.forename: Ke-feng
authors.initials: KF
authors.lastname: Dou
authors.forename: Zhi-nan
authors.initials: ZN
authors.lastname: Chen
mesh_terms: D000912:Antibodies, Neoplasm / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology*; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000276:immunology* / Q000473:pathology; D045744:Cell Line, Tumor; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology*; D008113:Liver Neoplasms / Q000276:immunology* / Q000473:pathology; D010843:Pichia / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology
chemical_list: D000912:Antibodies, Neoplasm; D007140:Immunoglobulin Fab Fragments; D011993:Recombinant Fusion Proteins