Reactive oxygen species (ROS) play crucial roles in both cell signaling and defense mechanisms. Hypochlorous acid (HOCl), a strong oxidant, aids the immune response by damaging pathogens. In this study, we developed two pyridinium-based fluorophores PSSM and PSSE for selective hypochlorite detection. Out of these two fluorescent probes, PSSM shows a strong turn-on emission via a photoinduced electron transfer (PeT) mechanism, excellent mitochondrial localization, and rapid response to HOCl with high selectivity among reactive oxygen species by achieving a detection limit of 2.41 μM. It successfully detects both exogenous and endogenous HOCl in live cells, enabling the study of HOCl's role at the organelle level. Structural analysis of PSSM via thioether oxidation confirmed by HPLC, NMR and HRMS further supports its specificity. Confocal imaging and flow cytometry studies further highlights its utility in investigating oxidative stress, positioning this fluorophore as a valuable tool for monitoring HOCl imbalances in biological systems.