Preparation and function assay of monoclonal antibodies against human CD20 by recombinant CD20 antigen
Objective To obtain monoclonal antibodies (McAb) against human CD20. Mothods BALB/c mice were immunized with CD20-expressing NIH-3T3 cells. The spleen cells of immunized mice were used to prepare the McAb by hybridoma techniques. CD20-McAb was selected by indirect immunofluorescence experiments on Daudi cells with cultural supernatant of hybridoma cells. Immunoprecipitation and flow cytometry (FCM) analysis were used to identify the surface molecules on Daudi cells recognized by the prepared antibody. Apoptosis assay and cell proliferation inhibition were examined by MTT, FCM and microscopy. Results A monoclonal antibody against CD20(1-28) was produced. The antibody recognized 33kD molecules on the surface of Daudi cells. FCM analysis indicated that 1-28 accorded with CD20-McAb in specific reaction with cell lines. Functional assay revealed that 1-28 could markedly inhibited the growth of Daudi cells and Raji cells, induced apoptosis of Daudi cells. Conclusion A hybridoma cell line 1-28 secreting CD20-McAb stably was obtained by recombinant CD20 antigen, and the CD20-McAb inhibited proliferation and induced apoptosis of CD20 positive cells.