title: Establishment of a monoclonal antibody against a peptide of the novel zinc finger protein ZNF32 proved to be specific and sensitive for immunological measurements.
abstract: BACKGROUND
  ZNF32 has been predicted to be a zinc finger protein and is involved in cell differentiation and tumor development, but its precise function is unknown. Specific monoclonal antibodies (mAbs) have been widely used in research and clinical diagnosis and treatments. Therefore, we established an anti-ZNF32 mAb to characterize this protein's function.
  
  
  METHODS
  Peptide⁴⁹⁻⁶³, a specific small peptide of ZNF32, was chosen and the synthetic keyhole limpet hemocyanin (KLH)-peptide⁴⁹⁻⁶³ was used as an antigen to immunize mice. A mAb against peptide⁴⁹⁻⁶³ was generated by hybridoma technology, and hybridoma cells were screened by limiting dilution. The isoform of mAb-pZNF32-8D9 was identified by double agar diffusion. The sensitivity and specificity of the mAb and expressed levels of ZNF32 in various cells and tissues were identified by enzyme-linked immunosorbent assay (ELISA), immunocytochemistry, immunohistochemistry, and Western blotting.
  
  
  RESULTS
  A stable anti-pZNF32-8D9 hybridoma secreting the anti-peptide49-63 mAb was established and the clone positive to the peptide⁴⁹⁻⁶³ in supernatant was 92% in ELISA. The mAb-pZNF32-8D9 is an immunoglobulin-1 that can be used for detecting the ZNF32 protein by immunocytochemistry, immunohistochemistry, and Western blotting and is highly sensitive and specific. We also found ZNF32 expressed at high levels in Jurkat and pulmonary squamous carcinoma cells, but it was not expressed in squamous epidermis cells.
  
  
  CONCLUSIONS
  mAb-pZNF32-8D9 can be used for the identification and expression of ZNF32. It might also provide a new tool for diagnostics or therapy for ZNF32-related diseases.
authors.affiliation: Division of Geriatrics, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, PR China.
authors.forename: Yuyan
authors.initials: Y
authors.lastname: Wei
authors.forename: Jie
authors.initials: J
authors.lastname: Zhang
authors.forename: Kai
authors.initials: K
authors.lastname: Li
authors.forename: Lugang
authors.initials: L
authors.lastname: Huang
authors.forename: Jun
authors.initials: J
authors.lastname: Li
authors.forename: Xiujie
authors.initials: X
authors.lastname: Wang
authors.forename: Ping
authors.initials: P
authors.lastname: Lin
authors.forename: Yuquan
authors.initials: Y
authors.lastname: Wei
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology; D015153:Blotting, Western; D002460:Cell Line; D002875:Chromosomes; D005260:Female; D006801:Humans; D006825:Hybridomas; D007150:Immunohistochemistry; D051741:Kruppel-Like Transcription Factors / Q000737:chemistry / Q000276:immunology*; D057230:Limit of Detection; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D051741:Kruppel-Like Transcription Factors; D010446:Peptide Fragments; C111477:ZNF32 protein, human