Preparation and biological character of monoclonal antibody against recombinant human epidermal growth factor

Objective To prepare high titer monoclonal antibodies against recombinant human epidermal growth factor (rhEGF). Methods The Balb/c mice were immunized with rhEGF and the spleen cells of Balb/c mice which were not immunized were used as feeder cells. Hybridoma technique and ELISA test were used to prepare and screen the hybridoma cell lines of monoclonal antibodies against rhEGF. The ascites developed by injecting the hybridoma cells into abdomal cavity of the Balb/c mice and purified with Protein A-Sepharose. The subclasses and isotypes were identified by mouse monoclonal antibody isotyping kit. The antigenic epitopes recognized by EGF-B 2, EGF-C 7, and EGF-A 8 were analyzed by the ELISA additivity test. Results Three hybridoma cell strains were obtained and named EGF-B 2, EGF-C 7, and EGF-A 8, which produced monoclonal antibodies specifically against rhEGF. The isotypes of the monoclonal antibodies were IgG 1κ, IgG 1λ, and IgG 3κ and the affinity constant were 2.76×10 10, 3.2×10 9, and 1.45×10 9, respectively. Conclusion We have successfully prepared three hybridoma cell strains which secrete high-titer and high-specific monoclonal antibodies against rhEGF. Our study provides a basis for the mechanism research and clinical applications of EGF and affords a valuable tool for EGF purification and tumor diagnose and treatment.