Specific Light-Up System for Protein and Metabolite Targets Triggered by Initiation Complex Formation

Abstract

Gene regulation systems are mimicked by simple quantitative detection of non-nucleic acid molecular targets such as protein and metabolite. Here, we describe a one-tube, one-step real-time quantitative detection methodology for isothermal signal amplification of those targets. Using this system, real-time quantitative detection of thrombin and streptomycin, which were used as examples for protein and metabolite targets, was successfully demonstrated with detection limits of at most 50 pM and 75 nM, respectively. Notably, the dynamic range of target concentrations could be obtained for over four orders of magnitude. Thus, our method is expected to serve as a point-of-care or on-site test for medical diagnosis and food and environmental hygiene.

Effects of Modifying Thioflavin T at the N3-Position on Its G4 Binding and Fluorescence Emission

“…We then demonstrated that ThT-HE binds to parallel G4 in particular and exhibits strong fluorescence while maintaining the low background emission properties of ThT [17]. In addition, we reported the application of ThT-HE to G4 detection systems [18][19][20]. Since then, ThT with an ethyl group introduced at the N 3 -position (ThT-E) has been reported by Guan et al [21], but the derivatization of ThT at the N 3 -position has not seen significant progress otherwise.…”

High-Contrast Facile Imaging with Target-Directing Fluorescent Molecular Rotors, the N3-Modified Thioflavin T Derivatives

“…Previously, we developed ThT derivatives by introducing substituents instead of methyl groups at the N 3 position adjacent to the rotational axis . We demonstrated that these fluorophores enabled the selective emission of fluorescence upon binding to parallel G4s. − However, so far, targets of ThT are still limited to β-sheet-rich proteins like amyloid fibrils and some nucleic acid structures mentioned above. Accordingly, in this study, we attempted to develop a fluorescence probe having a target-directing property that is different from that of the unmodified ThT in the conjugation of a ligand to the N 3 position.…”