Preparation and characterization of a novel monoclonal antibody specific to human EMILIN-5 protein

Hybridoma (Larchmt). 2007 Dec;26(6):417-21. doi: 10.1089/hyb.2007.0528.

Abstract

We aimed to establish the first monoclonal antibody (MAb) against the human EMILIN-5 protein (elastin microfibril interface located protein-5) and to investigate its distribution in normal human esophageal tissues and esophageal carcinomas. The bacterially expressed 6 His-EMILIN-5 fusion protein was induced and purified. Hybridomas were screened by indirect enzyme-linked immunosorbent assay (ELISA) using either purified 6x His-EMILIN-5 fusion proteins or purified 6x His-ZNRD1 fusion protein as a control. The EMILIN-5 protein-specific MAb was further identified by Western immunoblot analysis. The expression of EMILIN-5 was investigated in 63 cases of esophageal cancer specimens and 60 cases of normal esophageal specimens using immunohistochemical analysis. The expression of 6x His-EMILIN-5 fusion proteins was successfully induced. One MAb, H7 (IgG1), effective in detecting the recombinant and the cellular protein, was characterized. H7 bound to native EMILIN-5 protein and should be useful in studies of EMILIN-5 protein function and expression. EMILIN-5 staining was found positive in 37 cases of esophageal cancer tissues (59%) and 7 cases of normal esophageal tissues (12%). The higher-grade frequency of expression of EMILIN-5 in normal esophageal tissues was significantly lower (p < 0.01) than that in tumor tissues. We concluded that EMILIN-5 might play important roles in carcinogenesis of esophageal cancer.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aged
  • Animals
  • Antibodies, Monoclonal / biosynthesis*
  • Antibodies, Monoclonal / chemistry*
  • Antigens, Surface / chemistry
  • Antigens, Surface / genetics
  • Antigens, Surface / immunology*
  • Antigens, Surface / metabolism
  • Cell Line, Tumor
  • Esophageal Neoplasms / chemistry
  • Esophageal Neoplasms / etiology
  • Esophageal Neoplasms / metabolism
  • Female
  • Humans
  • Hybridomas
  • Male
  • Membrane Glycoproteins / chemistry
  • Membrane Glycoproteins / genetics
  • Membrane Glycoproteins / immunology*
  • Membrane Glycoproteins / metabolism
  • Mice
  • Mice, Inbred BALB C
  • Middle Aged
  • Neoplasm Proteins / chemistry
  • Neoplasm Proteins / immunology
  • Recombinant Fusion Proteins / immunology
  • Recombinant Fusion Proteins / isolation & purification
  • Recombinant Fusion Proteins / metabolism

Substances

  • Antibodies, Monoclonal
  • Antigens, Surface
  • EMILIN3 protein, human
  • Membrane Glycoproteins
  • Neoplasm Proteins
  • Recombinant Fusion Proteins
  • elastin microfibril interface located protein