Two-mode ratiometric electrochemical and turn-on fluorescence strategy for detecting biothiols in serum with all-in-one molecular probe.

Reactive sulfur species (RSS) play crucial roles in human physiology, with biothiols like hydrogen sulfide (HS), cysteine (Cys), and glutathione (GSH) being key regulators of oxidative stress and signal transduction. Here, a dual-mode resorufin-based (Re) small-molecule probe (Re-RSS) with electrochemical and fluorescent properties was developed for biothiol detection. Using a ratiometric electrochemical strategy and a "turn-on" fluorescence method, Re-RSS enabled sensitive quantification of HS, Cys, and GSH. The electrochemical approach showed linear ranges of 0-0.3 mM for HS, 0-0.1 mM for Cys, and 0-0.08 mM for GSH. Fluorescence spectrometry revealed linear relationships in the ranges of 1 μM-0.3 mM (HS), 2 μM-0.1 mM (Cys), and 5 μM-0.08 mM (GSH), with detection limits as low as 0.02, 0.01, and 0.08 μM (S/N = 3), respectively. This dual-mode sensor offers three major advantages: (1) dual-readout correction of system/background errors for improved accuracy; (2) independent signal transduction from distinct mechanisms, eliminating cross-interference; (3) negative-potential electrochemical detection free from biological interference. The direct dual-mode quantification highlights its significance and reliability for biothiol analysis in biological systems.