Minimal residual disease (MRD) monitoring has been demonstrated to important in predicting prognosis in acute myeloid leukemia (AML) receiving allogeneic hematopoietic stem cell transplantation (allo-HSCT), but the ideal time point and method remain unclear. Our study compared the prognostic value of multiparameter flow cytometry (MFC)-based and WT1 expression-based MRD a month before allo-HSCT [HSCT(-1 m)] and after allo-HSCT [HSCT(+ 1 m)], as well as next generation sequencing (NGS)-based MRD at HSCT(-1 m), HSCT(+ 1 m), 3 and 6 months after allo-HSCT [HSCT(+ 3 m) and HSCT(+ 6 m)] among 47 AML patients undergoing allo-HSCT. The MRD status by all the methods at HSCT(-1 m) was proved as a superior indicator with prognostic significance for disease progression, compared to that at HSCT(+ 1 m). For the NGS-based MRD, HSCT(+ 6 m) seemed to be the optimal detection time point, as supported by the optimal prognostic discrimination capability and the relatively high sensitivity for disease progression prediction. Moreover, our data showed that each individual method had some limitations in predicting prognosis; however, pre-transplant MRD monitoring by the combination of MFC, WT1 and NGS could greatly increase the sensitivity (100%) of identifying disease progression and greatly improve prognostic stratification. Our study may provide insights into the optimal time point and methodology for MRD monitoring in AML following allo-HSCT.