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To evaluate the relationship between myeloperoxidase (MPO) gene expression and specific lineages of hematopoietic differentiation, we analyzed expression of the MPO gene in a variety of normal and leukemic human cells. As a first step, we synthesized several oligonucleotide probes and isolated cDNA clones for the MPO gene. MPO expression was detected in most acute myeloid leukemias, and the level of expression correlated well with cytochemical enzymatic activity. No MPO message was detected in pure lymphoid leukemias, whereas very low basal levels of MPO transcripts were found in normal bone marrows. In four cases of acute undifferentiated leukemias cytochemically negative for MPO, significant levels of MPO transcripts were detected, suggesting a myeloid origin for these cases. These results indicate that MPO gene expression may serve as an additional marker for subclassification of acute leukemias and may be used to identify leukemic cells arrested at an early stage of the myeloid differentiation pathway.