Objective To investigate the effects of red orpiment on cell morphology, expression of promyelocytic leukemia (PML) mRNA and its protein localization in NB4 and HL-60 cell lines.Methods Cell morphology was assayed by Wright's staining and fluorescence staining, while PML mRNA expression was determined by RT-PCR. PML protein localization by evaluated by immunofluorescence staining. Results The typical apoptosis was found in NB4 and HL-60 cells after treatment with red orpiment. The fusion protein was no longer observed in NB4 cells, PML protein was relocated, and then degraded. In HL-60 cells, PML protein underwent a similar progress. The expression of promyelocytic leukemia (PML) mRNA was not changed in the treated cells.Conclusion Red orpiment inhibits the proliferation of leukemia cells by inducing them to undergo apoptosis.