Production of Monoclonal Antibody against Epstein-Barr Virus Early Antigen and Its Application to Detection of Antitumor Promoting Activity

摘要

Hybridoma cell line, designated 2D3-E7, producing a monoclonal antibody against 48 kDa-early antigen of Epstein-Barr vims was established by the fusion of mouse myeloma cells with spleen cells from a mouse immunized with P3HR-1 cells treated with sodium n-butyrate (1 mM) and a tumor promoter, 12-0-tetradecanoylphorbol-13-acetate (TPA, 40 ng/ml). The antibody was purified from the culture supernatant by precipitation with ammonium sulfate and column chromatographies on Sephadex G-2 00, Protein G-Sepharose, and Mono Q. Immunoblotting analysis showed that the purified antibody conjugated with horseradish peroxidase reacted with TPA (80 ng/ml)-induced polypeptide of 48 kDa in Raji cells and its induction was inhibited by curcumin (10 μ g/ml), an antitumor promoter from turmeric. These results indicated that the immunoblotting analysis can be used in a confirmation test for detection of antitumor promoting activity using the monoclonal antibody conjugated with the peroxidase. Furthermore, adhesion of Raji cells caused by treatment of TPA was inhibited by the curcumin in a dose-dependent manner, suggesting that the inhibition of adhesion can be also used in a preliminary test for detection of antitumor promoting activity.