We characterize B cell progenitors in the thymus. Although mature thymic B cells are surface Ig+, B220(high), but CD43-, B cell progenitors in the thymus were found to be slg-, B220(med), and CD43+. Most progenitors showed rearranged Ig D-J and V-D-J patterns when DNA from sorted B cell progenitors was amplified by PCR, blotted, and probed. When B cell progenitors were purified from the thymus and injected intrathymically into Ig heavy chain (Igh) allotype-disparate recipients, B cells bearing donor-type Igh6 were detected in the thymus but not in the periphery. Thymic B cells generated from these progenitors were CD5+ cells, although there was a broad range of expression of CD5 molecules. Furthermore, B cells purified from thymus that had been inoculated with B cell progenitors were able to differentiate into Ab-forming cells under the influence of CD40 ligand plus IL-10, and secreted IgM with donor Igh6 allotype. We thus clarified the existence of B cell progenitors in the thymus and also characterized the surface markers and their developmental functions.