Antigen receptor–mediated production of inositol-1,4,5-trisphosphate (Ins(1,4,5)P₃) in lymphocytes triggers the release of Ca²⁺ from intracellular stores; this release of Ca²⁺ results in the opening of store-operated Ca²⁺ channels in the plasma membrane. Here we report that mice lacking Ins(1,4,5)P₃ 3-kinase B (Itpkb), which converts Ins(1,4,5)P₃ to inositol-1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P₄), had impaired B lymphocyte development and defective immunoglobulin G3 antibody responses to a T lymphocyte–independent antigen. Itpkb-deficient B lymphocytes had the phenotypic and functional features of tolerant B lymphocytes and showed enhanced activity of store-operated Ca²⁺ channels after B lymphocyte receptor stimulation, which was reversed by the provision of exogenous Ins(1,3,4,5)P₄. Our data identify Itpkb and its product Ins(1,3,4,5)P₄ as inhibitors of store-operated Ca²⁺ channels and crucial regulators of B cell selection and activation.