A Label-Free Fluorometric Glutathione Assay Based on a Conformational Switch of G-quadruplex

In this paper, a label-free fluorescent method for glutathione (GSH) detection based on a thioflavin T/G-quadruplex conformational switch is developed. The sensing assay is fabricated depending on the virtue of mercury ions to form a thymine–thymine mismatch, which collapses the distance between two ssDNA and directs the guanine-rich part to form an intra-strand asymmetric split G-quadruplex. The newly formed G-quadruplex efficiently reacts with thioflavin T and enhances the fluorescent intensity. In the presence of GSH, Hg²⁺ is absorbed, destroying the G-quadruplex formation with a significant decrease in fluorescence emission. The proposed fluorescent assay exhibits a linear range between 0.03–5 μM of GSH with a detection limit of 9.8 nM. Furthermore, the efficacy of this method is examined using human serum samples to detect GSH. Besides GSH, other amino acids are also investigated in standard samples, which display satisfactory sensitivity and selectivity. Above all, we develop a method with features including potentiality, facility, sensitivity, and selectivity for analyzing GSH for clinical diagnostics.