Translocations involving chromosome band 11q23 are associated with acute lymphocytic and myelomonocytic leukemias with poor clinical prognosis. Pulsed-field gel electrophoresis (PFGE) was used to characterize the breakpoint region that has been mapped within a 300-kb fragment between the genes CD3G and PBGD. Using CD3G as a marker on SfuI-restricted DNA separated by PFGE, we detected a rearrangement involving 11q23 in the cell line B1 with a t(4;11) and in the leukemic cells of two patients, one with a t(2;11) and one with a t(11;19). In comparison, lymphoblastoid cell lines established from normal peripheral blood lymphocytes of these two patients had a normal karyotype and showed germline configuration, thus excluding RFL polymorphisms. Digestion of DNA with BssHII or SalI showed heterogeneity of 11q23 involving breakpoints. A rearrangement in the t(4;11) containing lymphoma cell line Karpas422 was seen only with the chromosome 4 probe KIT on SalI-digested DNA. PFGE is a reliable method for the mapping and detection of complex breakpoint regions. The breakpoints on 11q23 involve different introns of the highly spliced HRX/ALL-1/MLL gene.