Activity-Independent Enzyme-Powered Amplification for Improving Signal Stability and Fidelity in Biosensing

Enzymes are an important tool used for signal amplification in biosensing. However, traditional amplification methods based on enzymes are always dependent on their catalytic activities, so their signals fluctuate with the change of microenvironment (e.g., pH and temperature). In this work, we communicate an activity-independent enzyme-powered (AIEP) amplification strategy for biosensing to improve signal stability and fidelity. To verify this hypothesis, the monitoring of oxidative stress during drug-induced liver injury was carried out. Carboxylesterase (CEs), highly expressed in hepatic tissue, was selected as the amplification tool. A CEs configuration-matching fluorophore (CMF) was designed and screened, and a nanobeacon was fabricated by loading CMF within an O₂^•–-responsive polymeric micelle. Since the degradation of the nanobeacon was triggered by O₂^•–, CMF was released to bind with CEs, and the fluorescence was lit by CEs-CMF configuration matching but not catalytic reaction. Results demonstrated that the oxidative stress during drug-induced liver injury could be successfully monitored, and the hepatoprotective effects of repair drugs could be evaluated by cell and in vivo imaging. This strategy is flexible for bioactive molecules by altering the responsive unit and generally accessible for pharmacological evaluation.