Mitochondrial membrane potential (∆Ψm) plays central roles in cell apoptosis, signalling, metabolism, and other crucial bioevents. However, most of the current dual-emissive fluorescent probes for ∆Ψm works on aggregation mechanism, rendering possible interferences from inhomogeneous staining, cell numbers, and other factors. To resolve this knot, in this work we have developed a dual-emissive fluorescent probe detecting ∆Ψm based on polarity-response mechanism. The probe was designed to display different emission wavelength under different polarity, and simultaneously high affinity to both mitochondria and RNA. In live cells, mitochondria maintain a high negative transmembrane potential. Under this condition, the probe specifically targeted mitochondria because of its cationic structure. After the depolarization of ∆Ψm, the probe lost its mitochondrial targeting capability and shifted the target to RNA owing to its high RNA affinity. The distinct polar environments between the mitochondrial inner membrane and RNA result in a significant red shift in the fluorescence wavelength of the probe after its relocation from mitochondria to RNA. In this manner, the probe enabled the dual-emissive evaluation of the ∆Ψm. The probe was successfully applied to visualize the reversible change of ∆Ψm in real-time and in-situ manner. With the probe, the decrease of ∆Ψm in apoptosis procedure induced by toxins and anti-tumor drugs was also detected.