Two-color FACS analysis was used to study phenotypic subset and activation markers on circulating B (tumor) cells of 21 patients with chronic B-lymphocytic leukemia (B-CLL). Patients with clinically active (progressive) disease differed from patients with stable disease: B cells from the former patient category showed a significantly increased expression of the activation antigen 1D11 and FN99(CD9), and a decreased expression of the FN1 B subset marker. No clinical associations were observed using the CD23, CD25, 4F2, Ba, Bac-1 or FN50 markers. Functional studies showed that the B cells from both clinical categories of patients responded equally well with DNA synthesis when optimally triggered and supplied with T cell factors. However, B-CLL cells from patients with progressive disease secreted significantly higher levels of IgM in response to phorbol ester. The present experiments thus show that differences exist in the activation of B-CLL cells in vivo and that these patterns are correlated with disease activity. Further, the maximal in vitro proliferative capacity of individual tumor cells is similar, whereas differences in accessory T cell functions may exist between patients.