We used flow cytometry to evaluate constitutive PI3K-Akt-mTOR activation and the criteria for detection of two AML cell subpopulations were (i) two distinct and clearly separated cell subsets for one or more of the 18 mediators; and (ii) the smallest subpopulation being at least 20% of the total viable cell population. Here, AKT1 is linked to acute myeloid leukemia.