Objective：Based on the high affinity between Cu2+and sulfur atom in clinical commonly used anti-cancer drug 6-thioguanine（6-TG），a novel ratiometric fluorescence method was constructed for the detection of 6-TG. Methods：The o-phenylenediamine（OPD）could be oxidized by Cu2+ to its yellow fluorescence product 2，3-diaminophenazine（DAP）at 560 nm. The fluorescence intensity of carbon dot（CD）at 450 nm could be quenched by DAP because of the inner filter effect（IFE）. However，in the presence of 6-TG，it would compete with OPD for binding Cu2+，resulting in the reduction of the oxidation product DAP and its yellow fluorescence. Therefore，the IFE between the CD and DAP was weakened，and the blue fluorescence of the CD was restored. The fluorescence intensity ratio（F450/F560）was linearly correlated with the concentration of 6-TG. Therefore，this probe could be used to quantitatively detect 6-TG. Results：With the increase of 6-TG，the fluorescence intensity ratio（F450/F560）was gradually enhanced. Under the optimal reaction conditions，the linear concentration of 6-TG ranged from 40.00 to 160.00 μmol/L with detection limits of 61.10 nmol/L，and the correlation coefficient R2 was 0.992. Conclusion：This probe possesses good selectivity，high accuracy and is easy to operate. In addition，it is practical and economical，and can be applied to determine the content of 6-TG tablet.