Monoclonal antibody 9C4 recognizes epithelial cellular adhesion molecule, a cell surface antigen expressed in early steps of erythropoiesis

Exp Hematol. 2002 Jun;30(6):537-45. doi: 10.1016/s0301-472x(02)00798-1.


Objective: Monoclonal antibody (mAb) 9C4 detects a surface antigen expressed on immature erythroid progenitor cells and epithelial tumor cell lines. The aim of this study was to identify the recognized surface antigen and to analyze a potential role of this molecule in early steps of erythropoiesis.

Materials and methods: A pituitary-derived retroviral cDNA library was used to generate viruses and infect NIH-3T3 fibroblasts. The transfected cells were stained with mAb 9C4; positive cells were sorted by FACS; and a clonal cell line binding mAb 9C4 was established. cDNA encoding the 9C4-binding protein was amplified by polymerase chain reaction and cloned. Reactivity of mAb 9C4 with human bone marrow (BM) cells was analyzed by flow cytometry.

Results: Sequence analysis of the isolated cDNA uncovered a 100% identity with the epithelial cellular adhesion molecule (Ep-CAM). Two-color flow cytometric analysis revealed that almost 100% of Ep-CAM(+) BM cells coexpressed CD105, E-cadherin, and high levels of CD71. Fractions of Ep-CAM(+) BM cells also were CD34(+) but lacked glycophorin A expression, suggesting that Ep-CAM(+) cells represent immature erythroid cells. Reverse transcriptase polymerase chain reaction analysis of BM mononuclear cells revealed that the 9C4(+) erythroblast population but not the 9C4(-) fraction expressed Ep-CAM mRNA. Peripheral blood CD34(+) cells induced in vitro to differentiate into the erythroid lineage showed strong Ep-CAM expression on days 3 to 7 of culture. The addition of Ep-CAM-specific mAbs 9C4 or KS1/4 to the culture resulted in two- to three-fold up-regulation of Ep-CAM protein expression.

Conclusion: mAb 9C4 recognizes Ep-CAM, a molecule expressed in the early steps of erythropoiesis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3T3 Cells
  • Animals
  • Antibodies, Monoclonal*
  • Antibody Specificity
  • Antigens, Neoplasm / analysis*
  • Antigens, Neoplasm / genetics
  • Antigens, Surface / analysis*
  • Biomarkers, Tumor / analysis
  • Bone Marrow Cells / immunology
  • Cell Adhesion Molecules / analysis*
  • Cell Adhesion Molecules / genetics
  • DNA Primers
  • Epithelial Cell Adhesion Molecule
  • Erythropoiesis / physiology*
  • Flow Cytometry
  • Gene Library
  • Humans
  • Mice
  • Recombinant Proteins / analysis
  • Reverse Transcriptase Polymerase Chain Reaction
  • Transfection


  • Antibodies, Monoclonal
  • Antigens, Neoplasm
  • Antigens, Surface
  • Biomarkers, Tumor
  • Cell Adhesion Molecules
  • DNA Primers
  • Epithelial Cell Adhesion Molecule
  • Recombinant Proteins