Parkinson's disease (PD) is a neurodegenerative disorder caused by various factors such as neuroinflammation, oxidative stress, mitochondrial dysfunction, and neuronal apoptosis. Recent studies have shown that H2S supplementation reverses neuronal loss and mitigates motor deficits in PD patients through anti-inflammatory, antioxidant, improved mitochondrial function and proautophagic. Therefore, the discovery and use of H2S donors may be an exciting and intriguing strategy for the treatment of PD. Herein, we report a red emission mitochondria-targetable fluorescent probe, Rho-H2S, which can specifically and sensitively detect H2S with a limit of detection of 62.5 nM. Bioimaging experiments have shown that the probe has excellent mitochondrial targeting and good imaging capabilities for the detection of exogenous and endogenous H2S in cells. More importantly, based on the Rho-H2S probe, we first confirmed the sulforaphane (SFN) among 15 glucosinolate and isothiocyanate compounds from cruciferous vegetables with an outstanding ability to release H2S and we further proved that SFN could alleviate the symptoms of PD in vivo. All results demonstrate that Rho-H2S could be an effective tool for screening H2S donors and can contribute to the development of new therapeutic strategies for PD.