Reduced nicotinamide adenine dinucleotide (NADH) as a biomarker plays a crucial role in studying cell redox homeostasis. Simultaneously, the abnormal behavior of viscosity in cells is often accompanied by changes in the level of related active substances. Thus, real-time monitoring of in situ changes in NADH and viscosity is of great importance. Based on the specific recognition of NADH by the quinoline group, we developed a dual-responsive probe molecule (I-EQ), which is sensitive to both NADH and viscosity. The probe exhibits high sensitivity toward NADH, with a detection limit of 0.027 µM, and also shows excellent response to viscosity. Additionally, its application in monitoring NADH and viscosity in living cells was successfully demonstrated. Given these characteristics, I-EQ is expected to serve as a valuable dual-channel response tool for NADH and viscosity, with promising application in biomedicine and early diagnosis of diseases.