Although sulfur dioxide (SO2) plays an essential role in several physiological processes, monitoring of intracellular SO2 at subcellular levels remains challenging due to the lack of rapid and sensitive methods for its quantification in a 100% aqueous solution. Herein, a new hemicyanine dyes-based fluorescence probe, NBD-Id, was designed and synthesized for the detection of SO2 derivatives in pure aqueous solution and living cells. By virtue of a specific 1,4-addition reaction of SO3 2-HSO3 - and the polymethine chain of hemicyanine, significant changes in the absorption and fluorescence emission spectra were observed in less than 20 seconds. The ratiometric fluorescence (F467/F593) detection of SO2 derivatives was then obtained with high sensitivity (detection limit 3.6 nM). It was noted that NBD-Id has a specific response towards SO2 derivatives without interference from other anions and biomolecules. Intracellular fluorescence imaging indicated that NBD-Id is cell membrane permeable and mainly distributed within the mitochondria. Therefore, ratiometric fluorescence imaging of SO2 derivatives in the mitochondria of MCF-7 cells was successfully demonstrated.