The aim of this er vito study was to explore the potential of using the fluorescence lifetime of intraellular reduced nicotinamide adenine dinucleotide(phosphate)(NAD(P)H)as a label-free indicator to characterize the di ferencs between human leukemic myeloid cells and normal mononuclear cells(MNC).The steady-state and time-resolved autofuorescence of two human leukemic myeloid cell lines(K562,HL60)and MNC were measured by a spectrofuorimeter.According to excitation-enmission matrix(EEM)analysis,the optimal emission of NAD(P)H in these cells suspensions occurred at 445 nm.Furthermore,the fuorescence lifetimes of NAD(P)H in leukemic myeloid cells and MNC were determined by fitting the time-resolved autofuorescence data.The mean fuorescence lifetimes of NAD(P)H in K562,HL60,and MNC cells were 557±1.19,4.45±0.71,and 7.31±0.60 ns,respectively.There was a significant diference in the mean lifetime of NAD(P)H between leukemic myeloid cells and MNC(p<0.05).The difference was essentally caused by the change in relative concentration of free and protein-bound NAD(P)H.This study suggests that the mean fuorescence lifetime of NAD(P)H might be a potential label-free indicator for differentiating leukemic myeloid cells from MNC.