We have produced and characterized a new CD40 monoclonal antibody, mAb 89, which in the presence of anti-IgM antibodies co-stimulates to induce B cell proliferation. mAb 89 activates resting B cells as shown by an increase in cell volume and an enhanced subsequent proliferation of B cells in response to anti-IgM antibody. However, mAb 89 does not prepare B cells to respond to the growth-promoting activity of interleukin (IL) 2 or IL 4. Unlike IL 2 and IL 4, mAb 89 only weakly stimulates the proliferation of anti-IgM pre-activated B cells. Thus, the activating properties of anti-CD40 are likely to explain its co-stimulatory effect on B cells. Interestingly, the anti-CD40 mAb 89 was found to act in synergy with IL 4, but not with IL 2, in co-stimulation and restimulation assays. In this respect, anti-CD40 does not induce a significant increase of B cell surface IL 4 receptors while IL 4, but not IL 2, induces a twofold increase of the CD40 antigen expression. Thus the synergistic interaction between IL 4 and anti-CD40 may be related to the IL 4-dependent increase of CD40 antigen expression.