Preparation and Characterization of the Monoclonal Antibody against Chicken p15~(INK4B) Protein

Preparation of monoclonal antibody(McAb) against chicken p15INK4B(p15) protein was carried out for the function research of p15.The purified p15 protein as antigen immunized BALB/c mice.Two hybridoma,1C6 and 3F5,stably secreting McAb were produced by fusion of SP2/0 myeloma cells and spleen cells of syngeneic mice which had been previously immunized by p15 protein.Their titers in cell culture supernatant were 1∶820 and 1∶680 respectively,titers in ascites were 1∶4×106 and 1∶7×105 respectively,and affinity constants were 3.13×109 and 1.24×108 L/mol respectively.The monoclonal antibodies were of high affinity and different antigen binding epitope,and they could specifically bind to p15 protein purified from prokaryotic expressing product and expressed in BAC-TO-BAC Baculovirus Expression Systems but not other cell proteins.All of these show that the McAb against p15 was successfully prepared.