Src family kinases Fyn and Lyn associate with the B cell antigen receptor (BCR). Accumulating data show that Lyn plays important roles in BCR-mediated signaling, while the role of Fyn remains obscure. Here we dissected the role of Fyn and Lyn in BCR signaling using B cells from fyn^–/–, lyn^–/– and fyn/lyn double-deficient (fyn^–/–lyn^–/–) mice. In contrast to previous reports, fyn^–/– B cells were slightly hyporeactive to both anti-IgM and anti-IgD–dextran. Although lyn^–/– B cells were hyper-reactive to anti-IgM, anti-IgD-induced proliferation was impaired in lyn^–/– B cells. Most of the other phenotypes of fyn^–/–lyn^–/– mice were similar to that of lyn^–/– mice, except that proliferative responses of B cells to various stimuli, such as BCR cross-linking and lipopolysaccharide, were significantly lower in fyn^–/–lyn^–/– mice than in lyn^–/– mice. Finally, immune responses to thymus-independent type 2 antigen were affected in these mutant mice. These observations suggest that Fyn and Lyn are involved in B cell functions, and play similar, but partly distinct, roles in BCR signaling.