We reported the specific fluorescent probe (MC‐BOD‐XDS) with two‐steps reaction based on monosulfanyl‐coumarin‐BODIPY for selective detection of cysteine, high activity sulfanyl‐coumarin as the multiple reaction group instead of a group internal standard fluorophore. The reaction mechanism of MC‐BOD‐XDS for detecting cysteine was different from the reported probes about the nucleophilic aromatic substitution reaction (SNAr) of chlorinated BODIPY. The fluorescent color of MC‐BOD‐XDS changed from yellow to red, and then to orange. The linear calibration diagram showed that it can potentially be used for quantitatively detection of Cys. Its potential applications were demonstrated by employing it for detection of Cys in artificial urine and in fluorescent imaging in HeLa cells.
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