Enzyme-responsive aptasensing based on the ATP aptamer and benzothiazole-coumarin G-quadruplex fluorescent probe for label-free detection of adenosine deaminase activity and its inhibitor

Adenosine deaminase (ADA) is a vital enzyme for regulating the biological process in humans and its abnormal expression level is related to various severe diseases. Nevertheless, conventional approaches for detecting ADA activity are hindered by the drawbacks of complexity and limited detection performances. To address these challenges, we here designed an ADA-responsive aptasensing strategy based on a synthesized benzothiazole-coumarin G-quadruplex (G4) fluorescent probe, which can bind to the G-rich ATP aptamer (G-APA) and form the high-contrast fluorescent complex (S1/G-APA). While the fluorescence of S1/G-APA could be quenched by ATP competition due to the allosteric process, the deamination in ATP catalyzed by ADA would recover the fluorescence of S1/G-APA and allow a facile, turn-on, label-free, and selective detection of ADA activity. This proposed method exhibits favorable detection performances for ADA activity analysis with a wide detection linear range varied from 1 to 100 U/L and a detection limit of 0.82 U/L. Meanwhile, it can be applied for ADA inhibitor (DCF) evaluation and provide a turn-off and sensitive detection performances. Inspired by these detection efficacies, the serum samples spiked with ADA and DCF have been investigated to present the satisfied recovery rates and detection reliability. Collectively, this method will provide a promising way for portable and efficient ADA activity quantification and evaluating the inhibition efficiencies of ADA inhibitors, which hold the potential in promoting ADA-related sensing and pharmaceutical research.