title: [Protein expression, purification of PCGF1 and its monoclonal antibody preparation and identification].
abstract: OBJECTIVE
  To express the human recombinant PCGF1 protein and prepare monoclonal antibody (mAb) against it.
  
  
  METHODS
  The recombinant expression plasmid pET32a-His-PCGF1-128/189 was made and transformed into E.coli (BL21), and then the recombinant fusion protein His-PCGF1-128/189 was expressed and purified. The BALB/c mice were immuned with purified protein His-PCGF1-128/189 as antigen. The mAbs against PCGF1 were prepared by using standard hybridoma technique. The hybridoma cell lines were obtained by ELISA and Western blot screening procedure, the isotype of the mAbs were further identified by immune-double diffusion. Ascites were prepared from one propagated hybridoma cell line and mAbs were purified by using the Kit from Millipore. The valence of mAb was detected by Western blot.
  
  
  RESULTS
  The recombinant protein His-PCGF1-128/189 was expressed and purified. Two hybfidmas producing antibodies against PCGF1 were obtained, the isotypes of two mAbs were IgG1, Western blot showed that the antibodies were high sensitive(1:6 000) and high specific for PCGF1.
  
  
  CONCLUSIONS
  The anti-PCGF1 mAb prepared by using recombinant His-PCGF1-128/189 protein as antigen can be used for detecting PCGF1 proteins which are either endogenous or exogenous.
authors.affiliation: Department of Histology and Embryology, Medical College of Chinese People's Armed Police Force, Tianjin 300162, China. greylh@163.com
authors.forename: Hui
authors.initials: H
authors.lastname: Li
authors.forename: Xu-dong
authors.initials: XD
authors.lastname: Wu
authors.forename: Yan-hua
authors.initials: YH
authors.lastname: Gong
authors.forename: Bin
authors.initials: B
authors.lastname: Yin
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification; D015153:Blotting, Western / Q000379:methods; D045744:Cell Line, Tumor; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D005822:Genetic Vectors / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology / Q000302:isolation & purification; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids / Q000235:genetics; D063146:Polycomb-Group Proteins; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D012097:Repressor Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D063146:Polycomb-Group Proteins; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins