Approximately 5% of patients with chronic myelogenous leukemia (CML) do not reveal the Philadelphia (Ph) chromosome cytogenetically and are termed Ph-negative CML cases. We report one such case, which appeared normal by routine banding techniques. The BCR/ABL rearrangement was detected by reverse transcriptase-polymerase chain reaction and Southern blotting analysis, which suggested a b3-a2 splice junction. Dual color fluorescence in situ hybridization (FISH) with BCR and ABL DNA probes showed that the chimeric fusion gene was localized on chromosome 9q34, rather than at the typical location on chromosome 22q11. The BCR/ABL rearrangement was detected in 75% of the patient's bone-marrow population, whereas the remaining 25% of the cells appeared normal. The use of dual color FISH in the diagnosis of CML is extremely valuable not only in identifying cases of Ph-negative CML, but also in quantifying the proportion of transformed cell populations. This information ultimately results in an enhancement of our ability to monitor therapy, follow disease progression, and determine transplant eligibility.