Analysis of growth regulation in B-chronic lymphocytic leukemia (B-CLL) is of pivotal importance for understanding the pathophysiology and the development of new therapeutic approaches. We investigated the effect of soluble ligands and the interaction with fibroblasts in an in vitro system developed for the expansion of normal B lymphocytes. A total of 17 peripheral blood and bone marrow samples from patients with untreated B-CLL were analyzed for survival, apoptosis, and bcl-2 protein expression. The most efficient stimulus for cell survival was cocultivation with CDw32-transfected murine fibroblasts, which achieved a median of 56% surviving CD5 positive B cells with a plateau between Day 3 and Day 13 (p < 0.0001). IL-4 alone had a significant, but less profound, effect on cell survival: cell viability was increased by a factor of 1.7 on Day 3 (p = 0.001), but cell viability continued to decline. In contrast, the soluble recombinant human CD40 ligand and two different anti-CD40 antibodies did not prolong cell survival. In all experiments prolongation of cell survival was accompanied by a significant reduction of apoptosis of the leukemic B cells: in CDw32-transfected fibroblasts apoptosis was reduced by a mean of 90%, in IL-4 by a mean of 55%. Reduction in apoptotic cell death was associated with elevated bcl-2 protein levels. Our results emphasize the critical role of the interaction between B-CLL cells and CDw32-transfected fibroblasts for cell viability in vitro. Prolongation of cell survival is caused by a reduction of apoptosis and correlates with bcl-2 protein expression.