Objective To evaluate the value of flow cytometry ( FCM) for the differential diagnosis between myelodysplasia (MDS) subtype refractory cytopenia with multiple dysplasia (RCMD) and aplastic anemia (AA). Methods The flow cytometric data of bone marrow samples from 168 cases of RCMD and 77 cases of AA were analyzed retrospectively in blind, and its results were compared with gold standard to evaluate its diagnosis values. Results The specificity of abnormal of single immunophenotype in the surface of granulocytes and myeloblasts was high (range 75.3% -100% ) , but the sensitivity was very low (range 5.4%-50% ). In parallel tests, the sensitivity and specificity of the combination of CD+34 cells≥1% , myeloblasts ≥3% , abnormal expression of CD117 in granulocytes and loss of CD13 in myeloblasts or increased intensity of CD33 in granulocytes were higher than other combinations. The sensitivity and specificity of above combination were more than 62% and 92% , respectively. In the scoring method,different score was given to 8 markers according to different diagnostic value, which were CD+34 cells ≥1 % ,myeloblasts≥3% , abnormal expression of CD117 in granulocytes, loss of CD13 in myeloblasts, increased intensity of CD33 in granulocytes, loss of CD13 in granulocytes, loss of CD10 in granulocytes, and decreased SSC in granulocytes. The sensitivity and specificity were both high if we defined that the total score ≥1.5 was RCMD and the score < 1. 5 was AA. Conclusions The value of abnormal of single immunophenotype for differential diagnosis between RCMD and AA is low. Parallel tests can increase the diagnostic sensitivity obviously and not decrease the specificity. CD+34 cells≥1% , myeloblasts≥3% and abnormal expression of CO117 in granulocytes were the most important markers. The scoring method is precise to distinguish RCMD from AA.