Preparation And Functional Study Of Monoclonal Antibodies Against The Human CD20 And CD34

At present, the monoclonal antibodies (McAbs) against the molecules on the cell surface are being used in clinics for the diagnosis and treatment of various diseases Anti-CD20 McAb have been successfully employed in the clinical treatment of B-cell lymphomas in both unmodified and radiolabeled forms Clinical studies of anti-CD20 McAb have demonstrated considerable anti-tumor activity and safety The CD34 McAb is now commonly used for enrichment of hemopoietic progenitors for bone marrow transplantation In the present study, the recombinant antigens were expressed by NIH3T3 cells either transfected with the plasmids containing CD20 cDNA or infected with the recombinant vaccinia virus containing CD34 cDNA to obtain the anti-CD20 and anti-CD34 McAbs, which are being widely used, but not commercially made in China The anti-CD20 McAb named 1-28 was identified by indirect immunofluorescence with Daudi cells The specificity of 1-28 was determined by immunoprecipitation and flow cytometry assay (FCM) The results showed that 33kD molecules on the surface of Daudi cells were recognized by 1-28 The specific reaction with different cell lines was tested to be accorded with standard anti-CD20 McAb 1-28 could compete with standard anti-CD20 McAb to bind to CD20 molecules on the cytoplasmic membrane of Daudi cells The growth inhibition and apoptosis induced by 1-28 were confirmed by several different assays, including MTT or trypan blue staining, PI staining in FCM, electron and optical microscopy The signaling events involved in anti-CD20-induced apoptosis were investigated Increased intracellular Ca2+ concentration, caspase activation, and cleavage of caspase substrates were observed The results also indicated that complement-dependent cytotoxicity (CDC) was another important effector mechanism in Daudi cell killing 1,25-(OH)2VD3 was found to inhibit proliferation of human myeloma cell line RPMI8226 and stimulate the expression of CD20 molecules on the cytoplasmic membrane of RPMI8226 cells 1-28 was proved to have a cooperative effect on growth inhibition of the RPMI8226 cells by 1,25-(OH)2VD3 Further studies of theanti-tumor effect of 1-28 should provide new insights into more effective therapies for B-cell lymphomas Two hybridoma cell lines 5D41C12 and 18-15 secreting anti-CD34 McAb were selected by indirect immunofluorescence with KG1-a cells However, 18-15 turned negative to KG1-a cells later The specificity of 5D41C12 are in research The preparation of anti-CD34 McAb is important not noly for autologous marrow reinfusion, but also for the exploration of the regulation and function of CD34 molecules...