title: [Preparation and characterization of a polyclonal antibody against mouse NH2;-terminal domain of polycystin-1].
abstract: OBJECTIVE
  To produce a rabbit polyclonal antibody, mPkd1-Np, against the extracellular portions of polycystin-1 (PC1) in order to explore the functional roles of the PC1 NH2;-terminus.
  
  
  METHODS
  Based on hydrophobic/hydrophilic analyses, we chose a cDNA fragment that encodes amino acids 474E-640L on PC1 and amplified it via RT-PCR. The PCR product was then cloned into a prokaryotic expression vector pGEX-GST. After IPTG induction, the antigen mPkd1-N was produced and further purified. A rabbit was immunized with this antigen and its antiserum was collected. The mPkd1-Np antibody was validated to be specific for PC1 protein through Western blotting, immunohistochemistry, and immunofluorescence methods.
  
  
  RESULTS
  The prokaryotic expression vector pGEX-mPkd1-N was successfully constructed and mPkd1-N antigen was induced to express in E.coli Rossetta cells. Using this antigen, the polyclonal antibody mPkd1-Np was produced and its specificity for PC1 was proved through biochemistry and cellular assays.
  
  
  CONCLUSIONS
  We successfully produced an anti-PC1 NH2;-terminal polyclonal antibody named mPkd1-Np. The polyclonal antibody provides a platform for further research into PC1 NH2;-terminal function, specifically renal tubulogenesis and its maintenance.
authors.affiliation: Division of Translational Cancer Research and Therapy, State Key Laboratory of Molecular Oncology, Cancer Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100021, China.
authors.forename: Haichao
authors.initials: H
authors.lastname: Liu
authors.forename: Yuan
authors.initials: Y
authors.lastname: Li
authors.forename: Ao
authors.initials: A
authors.lastname: Li
authors.forename: Yuan
authors.initials: Y
authors.lastname: Chen
authors.forename: Chunyang
authors.initials: C
authors.lastname: Sun
authors.forename: Zhenwei
authors.initials: Z
authors.lastname: Ding
authors.forename: Guanqing
authors.initials: G
authors.lastname: Wu
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology* / Q000302:isolation & purification*; D000918:Antibody Specificity / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D023061:Gene Order; D005822:Genetic Vectors / Q000235:genetics; D007668:Kidney / Q000378:metabolism; D051379:Mice; D016366:Open Reading Frames; D054730:Protein Interaction Domains and Motifs / Q000235:genetics / Q000276:immunology*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D050396:TRPP Cation Channels / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
chemical_list: D000906:Antibodies; D011993:Recombinant Fusion Proteins; D050396:TRPP Cation Channels