Rearrangements within the chromosome 11q13 region are frequent in hematologic malignancies. 50% of 75% of mantle cell lymphomas (MCLs) carry a translocation t(11;14) (q13;q32). Using Southern blot analysis, a BCL1 breakpoint can be detected in approximately 50% of MCLs. It is not known whether other MCLs harbor also breakpoints at 11q13. Breakpoints in this region not involved in t(11;14), are detected in chronic lymphocytic leukemia and acute myeloid leukemia. To detect and localize breakpoints at 11q13 more accurately, we have developed fluorescence in situ hybridization using two probe sets of differently labeled cosmids, symmetrically localized at either side of the major translocation cluster of BCL1. These probes span a region of 450 to 750 kb. We applied this assay to a series of hematologic malignancies with 11q13 abnormalities identified by classical cytogenetics. All four samples with a t(11;14) (q13;q32) showed dissociation of the differently colored signals in metaphase and interphase cells, thereby indicating a chromosomal break in the region defined by the probe sets. The frequency of abnormal metaphase and interphase cells was comparable with that observed in any of the 13 malignancies with other chromosomal 11q13 abnormalities, indicating that these chromosomal breaks occurred outside the 450- to 750-kb region covered by the probes. One patient showed triplication and one patient showed monoallelic loss of this region. The current data show that double-color fluorescence in situ hybridization is a simple and reliable method for detection of the t(11;14)(q13;q32) in interphase cell nuclei and that is can be used to distinguish this translocation from other 11q13 rearrangements in hematologic malignancies.