Expression of CD40 on mouse cells was investigated comparing the binding to cells of a monoclonal antibody against CD40, to that of a soluble fusion protein consisting of the extracellular domains of the mouse CD40‐ligand. The analysis of a series ofestablished cell lines failed to demonstrate expression of CD40 on pro‐ or pre‐B cells, and indicated that CD40 expression was restricted to cells that had undergone productive heavy‐ and light‐chain gene rearrangements, and expressed surface Ig. In cells from normal mice, CD40 first becomes detectable, although at low levels, on a subset of small pre‐B‐II cells in bone marrow, the levels of CD40 expression increasing thereafter during B‐cell maturation. Thus, immature B cells (IgM+IgDloB220lo) express intermediate levels of CD40, and mature B cells (IgM+IgDhiB220hi) express high levels of CD40. Anatomical location also seems to correlate with the levels of CD40expression, as B cells expressing the highest levels of CD40 were found in lymph nodes and