Although evidence suggests that peripheral B cell death plays a role in maintenance of B cell homeostasis including elimination of self-reactive B cells, the site of B cell death in unimmunized peripheral lymphoid organs has not yet been studied. Here we demonstrate that cells exhibiting reduced mitochondrial membrane potential and those expressing active caspase 3, both of which are characteristic for apoptosis, are specifically accumulated in the marginal zone (MZ) B cell compartment compared to the other B cell subsets in normal unimmunized mouse spleen. These apoptotic MZ B cells exhibit increased expression of MHC class II and CD22, whose expression is shown to be increased upon B cell activation, but not other activation markers such as CD69 and CD86, suggesting that apoptotic MZ B cells are activated with partial expression of B-cell-activation-associated antigens. These results indicate that MZ is the major site of peripheral B cell apoptosis in unimmunized mice. Apoptosis of MHC class II(hi) CD22(hi) MZ B cells may contribute to B cell homeostasis.