Hypochlorite (ClO−) and viscosity both affect the physiological state of mitochondria, and their abnormal levels are closely related to many common diseases. Therefore, it is of great significance to develop a mitochondria-targeting fluorescent probes for the dual sensing of ClO− and viscosity. Herein, we have explored a new fluorescent probe XTAP−Bn, which responds sensitively to ClO− and viscosity with off-on fluorescence changes at 558 and 765 nm, respectively. Because the emission wavelength gap is more than 200 nm, XTAP−Bn can effectively eliminate the signal crosstalk during the simultaneous detection of ClO− and viscosity. In addition, XTAP−Bn exhibits a series of advantages, including high selectivity, rapid response, good water solubility, low cytotoxicity, and excellent mitochondrial targeting ability. More importantly, probe XTAP−Bn has been successfully employed to monitor the dynamic change of ClO− and viscosity levels in mitochondria of living cells and zebrafish. Taken together, this work not only provides a reliable tool for identifying mitochondrial dysfunction, but also offers a potential approach for the early diagnosis of mitochondrial-related diseases.