Fluorescence Biosensing Based on Bifurcated DNA Scaffold-Aggregated Ag Nanocluster via Responsive Conformation Switch of Quasi-Molecular Beacon.

To address the limitations of typical hairpin-structural molecular beacons, exploring the ability of a quasi-molecular beacon ( q MB) to create label-free fluorescence biosensors is intriguing and remains a challenge. Herein, we propose the first example of modular q MB with the feature of a stimulation-responsive conformation switch to develop an aggregated Ag nanocluster ( a AgNC) in a bifurcated DNA scaffold for fluorescently sensing a specific initiator ( I *). This q MB was well designed to program four functional modules: I *-recognizable element adopting metastable stem-loop bihairpin structure and two DNA splits (exposed C 3 GT 4 and locked C 4 AC 4 T) of a AgNC template that is separated by a tunable hairpin spacer for the customized combination of selective recognition and signaling readout. When presenting I * in an assay route, the specific hybridization induces the directional disassembly of the bihairpin unit, on which the q MB is configurationally switched to liberate the locked split. Thus, the bifurcated parent template pair of C 3 GT 4 /C 4 AC 4 T is proximal, affording in situ nucleation and clustering of emissive a AgNC. By collecting the fluorescence signal, the quantitative detection of I * is achieved. Benefiting from the ingenious programming of q MB, the recognizing and signaling integration actuates the construction of a facile and convenient fluorescent biosensor featuring rapid reaction kinetics, a wide linear range, high sensitivity, and specificity. This would provide a new paradigm to exploit versatile q MB-based biosensing platforms via stimulation-responsive conformation switches for developing various DNA-scaffolded Ag clusters.