A novel label-free ratiometric fluorescent probe is developed for the detection of H 2 O 2 based on GelRed/[G 3 T] 5 /Tb 3+ hybrid, in which GelRed (a nucleic acid dye) intercalated into the designed single-stranded DNA [G 3 T] 5 (i.e. GelRed/[G 3 T] 5 ) acts as a stable build-in reference with red emission and Tb 3+ sensitized by [G 3 T] 5 (i.e. [G 3 T] 5 /Tb 3+ ) as a sensitive response signal with green fluorescence. With the successive addition of Hg 2+ and cysteine (Cys) to GelRed/[G 3 T] 5 /Tb 3+ , the fluorescence of [G 3 T] 5 /Tb 3+ can be effectively quenched and recovered respectively, while the fluorescence of GelRed/[G 3 T] 5 remains unchangeable. Combined with these properties, we have demonstrated its application for label-free ratiometric fluorescence detection of H 2 O 2 with self-calibration. The sensing mechanism is based on the specific reaction between H 2 O 2 and Cys, the resulting disulfide reverses the Cys-mediated fluorescence changes of [G 3 T] 5 /Tb 3+ . This method is further applied to the monitoring of oxidase-related reactions. As the glucose oxidase (GOx)-biocatalyzed oxidation of glucose and the acetylcholine esterase/choline oxidase (AChE/ChOx) cascade yield H 2 O 2 , such two biocatalytic processes are successfully examined utilized our proposed method, indicating its potential broad applications in biomedical analysis. In addition, we have also demonstrated its feasibility for oxidase-based visual biosensing, in which only a handheld UV lamp is used. Importantly, coupled with our proposed visual biosensing, a realtime quantitative scanometric assays has been also demonstrated by using a smartphone with easy-to-access color-scanning APP as the detection platform. Compared to traditional methods, the proposed design is cost-effective, simple to prepare and easy-to-use without fluorescence labeling or chemical modification.
GelRed-photosensitized DNA cleavage enabled Hg 2+ detection. • T-Hg 2+ -T coordination triggered dsDNA assembly and GR intercalation. • The biosensor showed a linear range of 0.01–100 nM and an LOD of 1.30 pM. • Spiked water samples gave recoveries of 83.89–119.43%. A label-free GelRed-photosensitized electrochemical DNA biosensor was developed for Hg 2+ detection in water based on light-induced DNA cleavage. The sensing strategy integrates Hg 2+ -specific T-Hg 2+ -T coordination, GelRed (GR) intercalation, photosensitized oxidation, and impedance-based electrochemical readout. In the presence of Hg 2+ , two thymine-rich single-stranded DNA probes assemble into a double-stranded DNA structure through T-Hg 2+ -T coordination, allowing the nucleic acid dye GR to intercalate into the double-stranded DNA (dsDNA). Upon green-light irradiation, the dsDNA-GR complex acts as a photosensitizing system to generate singlet oxygen ( 1 O 2 ), which induces oxidative cleavage of the DNA and consequently changes the interfacial R ct . Under optimized conditions, R ct showed a linear relationship with the logarithm of Hg 2+ concentration, described by R ct (Ω) = −907 log C (Hg 2+ ) + 5312. The biosensor exhibited a linear range of 0.01–100 nM and a low limit of detection (LOD) of 1.30 pM. In spiked environmental water samples, recoveries of 83.89–119.43% were obtained with relative standard deviations of 3.49–9.32%, and the results were further evaluated by comparison with atomic fluorescence spectrometry. This work provides a GelRed-photosensitized DNA-cleavage strategy for sensitive and label-free electrochemical monitoring of Hg 2+ in aquatic environments. A label-free photocatalytic electrochemical DNA biosensor was developed for the detection of Hg 2+ in water. Download: Download high-res image (126KB) Download: Download full-size image
危险品化学品经营许可证（带存储）