A 47-year-old man was admitted to our hospital in June 2009 because of fatigue and blast cells in peripheral blood. Bone marrow examination showed that 67% leukemic cells were positive for myeloperoxidase (MPO) and negative for esterase stain. Flow cytometric analysis (FCM) revealed the expressions of CD2, cyCD3, CD5, TdT, CD13 on the blasts. Chromosome analysis of the bone marrow cells demonstrated 46, XY in 18 of the 20 analyzed cells, 46,XY,t(1;11)(q21;p15) in 1 of the 20 analyzed cells, and 47,XY,+11 in 1 of the 20 analyzed cells. The patient was diagnosed as having mixed phenotype acute leukemia, T/myeloid, NOS, according to the WHO classification. He received induction chemotherapy for ALL, but could not achieve complete remission (CR). After initial treatment, residual leukemic cells with CD13, CD33 and MPO were detected by FCM; therefore, he received re-induction chemotherapy for AML, and achieved CR. Acute leukemia of ambiguous lineage is a relatively rare subtype in acute leukemia and standard chemotherapy has not been established. It was suggested that the selection of chemotherapy based on the results of FCM was successful in our patient.