In the pathogenesis of microglia, brain immune cells promote nitrergic stress by overproducing nitric oxide (NO), leading to neuroinflammation. Furthermore, NO has been linked to COVID-19 progression, which has caused significant morbidity and mortality. SARS-CoV-2 infection activates inflammation by releasing excess NO and causing cell death in human microglial clone 3 (HMC3). In addition, NO regulates lysosomal functions and complex machinery to neutralize pathogens through phagocytosis. Therefore, developing lysosome-specific NO probes to monitor phagocytosis in microglia during the COVID-19 infection would be a significant study. Herein, a unique synthetic strategy was adopted to develop a NO selective fluorescent probe, , which can discriminate activated microglia from their resting state. The nonfluorescent exhibits a turn-on response toward NO only at lysosomal pH (4.5-5.5). Quantum chemical calculations (DFT/TD-DFT/PCM) and photophysical study revealed that the photoinduced electron transfer (PET) process is pivotal in tuning optical properties. demonstrated good biocompatibility and lysosomal specificity in activated HMC3 cells. Moreover, it can effectively map the dynamics of lysosomal NO against SARS-CoV-2 RNA-induced neuroinflammation in HMC3. Thus, is a potential fluorescent marker for detecting RNA virus infection and monitoring phagocytosis in HMC3.