Richter's syndrome is defined as the morphologic transformation of chronic lymphocytic leukemia (CLL) into diffuse large-cell lymphoma (DLL). To determine the clonal nature of the two malignancies, we microdissected the CLL and DLL cells from a lymph node of Richter's syndrome and analyzed the sequences of the rearranged Ig VH-D-JH genes of the two lymphomas. Using the Ig VH-D-JH sequence as a marker of clonality, we delineated the clonal relationship of the CLL and DLL cells. The microdissected CLL and DLL cells productively rearranged different VH, D, and JH genes, suggesting that these DLL B cells emerge as discrete elements independent of the CLL B-cell population. The productively rearranged Ig V gene sequence of the CLL clone was 100% identical to the VH6 germline gene, but the rearranged Ig VH gene of the DLL clone was somatically point-mutated based on comparison of its sequence with those of reported germline genes. In the DLL clone, the random distribution and nature of the somatic point-mutations suggests a lack of antigen selection; the identity of the somatic point-mutations in multiple independent isolates of the same B-cell clone suggests a lack of intraclonal diversity. Thus, Richter's syndrome DLL B cells are monoclonal and can emerge as discrete elements independent of the preexisting CLL cells; antigen selection and clonal diversification are not necessarily associated with the events leading to this aggressive neoplastic transformation.