Characterization of monoclonal antibody CIBCNSH3 generated to the human EGF receptor

Monoclonal antibody CIBCNSH3 of IgG1 isotype has been generated against human epidermal growth factor receptor (EGFR) using MDA MB 468 breast carcinoma cell line as immunogen. Earlier studies have revealed that this MAb blocked growth factor-receptor interaction and thus inhibited cell proliferation and tumor growth. In the present paper, this MAb has been extensively characterized to evaluate its application in the study of human cancers. The results were compared with those obtained using a control MAb ICR 62 specific to EGFR. Competitive assay showed that this MAb bound to an epitope in the extracellular domain of the EGFR to which MAb ICR 62 also bound. This MAb immunoprecipitated the 170 kD glycoprotein. The specificity was further confirmed by the formation of a single discrete band in western blot analysis. By flow cytometric analysis this monoclonal antibody revealed high binding affinity with MDA MB 468 cells. By immunocytochemical assay, out of 35 breast tumors studied, 40% were found to exhibit strong cell membrane staining and in the case of 25 oral cancers studied, 56% were strong positive. High expression of EGFR was observed in MDA MB 468 cells and HN 5 cells. These studies clearly indicate that MAb CIBCNSH3 might prove useful to identify tumors with high level of expression of EGFR associated with poor prognosis.