BACKGROUND: Leukemia stem cells (LSCs) in play an important role in the initiation, relapse, and progression of acute myeloid leukemia (AML), and in the development of chemotherapeutic drug resistance in AML. Studies regarding the detection of LSCs and the development of novel therapies for targeting them are extensive. The identification of LSCs and targeting therapies for them has been continuously under investigation. METHODS: We examined the levels of CD45CD34CD38CD133 cells in bone marrow samples from patients with hematological malignancies and healthy controls, using four-color flow cytometry. RESULTS: Interestingly, the CD45CD34CD38CD133 cells were highly expressed in the bone marrow of patients with AML compared to that in healthy controls (HC). Moreover, the proportions of CD45CD34CD38CD133 cells were also examined in diverse hematological malignancies, including AML, CML, DLBCL, MM, MDS, HL, ALL, and CLL. LSCs were prominently detected in the BMCs isolated from patients with AML and CML, but rarely in BMCs isolated from patients with DLBCL, MM, MDS, ALL, CLL, and HL. Additionally, the high CD45CD34CD38CD133 cell counts in AML patients served as a significantly poor risk factor for overall and event free survival. CONCLUSIONS: Therefore, our results suggest that CD45CD34CD38CD133 cells in AML might potentially serve as LSCs. In addition, this cell population might represent a novel therapeutic target in AML.