A Hg2+-mediated label-free fluorescent sensing strategy based on G-quadruplex formation for selective detection of glutathione and cysteine

Abstract

A novel label-free fluorescent strategy for the detection of glutathione (GSH) and cysteine (Cys) is presented. The system consists of two single stranded DNA (ssDNA) with thymine–thymine (T–T) mismatches and used Hg²⁺ as a mediator, and N-methyl mesoporphyrin IX (NMM) as the signal reporter. The assay is based on the competitive reaction of Hg²⁺ with GSH/Cys and T–T mismatched double stranded DNA (dsDNA). In the absence of the target, two ssDNA containing T–T mismatches react with Hg²⁺ to form a T–Hg²⁺–T dsDNA structure in the solution, which hampers the formation of a G-quadruplex structure. However, in the presence of the target, GSH/Cys reacts with Hg²⁺ to keep DNA probes in a free single state, resulting in the effective formation of a G-quadruplex structure of the DNA probe (GP). Subsequently, due to the strong interaction between the G-quadruplex structure and NMM, fluorescence was greatly enhanced. This fluorescence strategy does not require any chemical modification, making the assay convenient and cost-effective. This method exhibited a linear relationship between peak fluorescence intensity and concentration of GSH in the range of 10–400 nM with a limit of detection (LOD) of 9.6 nM. A linear range for Cys detection was obtained in the concentration range of 10–500 nM with an LOD of 10 nM. Moreover, the proposed method worked well for the analysis of complex biological samples.