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Hydrogen sulfide (H 2 S) is a multifunctional gaseous signaling molecule that plays a vital role in several biological processes. In the present study, a BODIPY-based fluorescent probe called 8-[4-((1,4,7,10-tetraazacyclododecane)methyl)phenyl]-4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a, 4a-diaza- s -indacene (BA–Cyclen)–Cu was designed and synthesized; this probe is a Cu(Ⅱ) complex that uses Cu(Ⅱ) decomplexation to achieve the sensitive and rapid detection of aqueous H 2 S via the "turn-on" mode. We observed that BA–Cyclen–Cu exhibited good membrane permeability, low toxicity, and lysosome-targeting ability, facilitating H 2 S detection in living cells. Furthermore, we demonstrated the potential biological applications of the probe by measuring exogenous H 2 S originating from Na 2 S and GYY4137, a slow-release donor, and endogenous H 2 S generated via the catalysis of cystathionine- β -synthase in both normal (H9c2) and cancerous (U87) cells. Moreover, BA–Cyclen–Cu was successfully used to detect exogenous H 2 S by the external standard method in fetal bovine serum, the serum of a healthy person, and the serum of a patient with liver cancer. [Display omitted] • BA-Cyclen-Cu displayed high selectivity, sensitivity and fast response toward H 2 S with "turn-on" mode. • BA-Cyclen-Cu have a good performance on exogenous/endogenous H 2 S sensing in cancer cells and normal cells. • BA-Cyclen-Cu was successfully used to detect H 2 S in serum samples. [ABSTRACT FROM AUTHOR]