A Reciprocal-Amplifiable Fluorescence Sensing Platform via Replicated Hybridization Chain Reaction for Hosting Concatenated Multi-Ag Nanoclusters as Signal Reporter

Abstract

Exploring the replication of hybridization chain reaction HCR (rHCR) for reciprocal amplification is intriguing in biosensing and bioanalysis. Herein, we develop a rHCR-based fluorescence platform that is manipulated by the combination of a specific DNA trigger ( T ) and a T -analogous amplicon ( T* ), thereby concatenating multi green-emissive Ag nanoclusters (mgAgNCs) for amplifiable signal readout. Four well-designed hairpins (H1 recognizing T , H2, H3, and H4) with sequential complements are executed to operate rHCR. The termini of H1/H3 are merged to hybridize an inhibiting strand (I). The parent scaffold for mgAgNCs is separated into two splits (C4AC4T and C3GT4) that are individually overhung in H2/H4. The presence of T activates the first HCR amplifier through cross-hybridization of four reactive hairpins for forming HCR duplexes. The next invasion of a complex ( T* ·I) drives I to hybridize the tandem repeats in H1/H3, so that the displaced T* functions as T to catalyze the second amplifier rHCR for feeding back more hairpin assemblies with rapid reaction kinetics. In the shared rHCR polymers, the parent scaffolds (C4AC4TC3GT4) in H2/H4 are collectively concatenated for the preferential clustering of mgAgNCs adducts, which cooperatively emit enormous T -responsive fluorescence signal. Because of the localization of T in HCR products, an alternative amplicon T* is introduced to drive rHCR progress via DNA strand displacement, generating more nucleating sites of emitters. Thus, the rational combination of nonenzymatic rHCR and label-free fluorescent concatemers would create a reciprocal signal amplification, achieving a simplified, rapid, and highly sensitive assay down to femtomolar concentrations.

AgAuS Quantum Dots as a Highly Efficient Near-Infrared Electrochemiluminescence Emitter for the Ultrasensitive Detection of MicroRNA

“…11,12 Therefore, the rapid and accurate detection of trace miRNA is particularly important to achieve early disease diagnosis. Over the years, nucleic acid amplification strategies (rolling circle amplification, 13 hybridization chain reaction, 14 catalytic hairpin assembly, 15 etc.) had been proposed and applied in the fabrication of biosensor for miRNA detection.…”

“…MicroRNA-222 (miRNA-222) played a critical role as a regulator of gene expression in some diseases such as liver cancer and rheumatoid arthritis, , whose expression was associated with toxic heavy metal exposure in the environment. , Therefore, the rapid and accurate detection of trace miRNA is particularly important to achieve early disease diagnosis. Over the years, nucleic acid amplification strategies (rolling circle amplification, hybridization chain reaction, catalytic hairpin assembly, etc.) had been proposed and applied in the fabrication of biosensor for miRNA detection.…”

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

“…For example, the bifurcated fragments of the AgNC parent template can be easily combined via multiple interconversions of conformations or orientated self-assembly of DNA nanostructures upon stimulation by an initiator. − Moreover, Petty’s group conducted an interesting study that focused on short duplexes locally concentrating two split templates by directional DNA assemblies . In our group, we have demonstrated the highly stable fluorescent performances and unique spectral behavior of green AgNC by separating the template into two splits and encoding them in the tails of hairpins, pointing to the construction of fluorescence biosensing platforms. , These progress would offer great possibilities to exploit the programming of AgNC template splits in noncanonical DNA components for label-free assay.…”

“…27 In our group, we have demonstrated the highly stable fluorescent performances and unique spectral behavior of green AgNC by separating the template into two splits and encoding them in the tails of hairpins, pointing to the construction of fluorescence biosensing platforms. 28,29 These progress would offer great possibilities to exploit the programming of AgNC template splits in noncanonical DNA components for label-free assay.…”

Methylation-Powered Assembly of a Single Quantum Dot-Based FRET Nanosensor for Antibody-Free and Enzyme-Free Monitoring of Locus-Specific N6-Methyladenosine in Clinical Tissues

“…Despite their improved sensitivity, more attention should be paid to coordinating various parameters (e.g., temperature, pH, ionic concentration, and strength) to avoid false positives caused by undesired enzymic polymerization. Programmable enzyme-free amplification circuits (e.g., hybridization chain reaction (HCR) and catalytic hairpin assembly (CHA)) can be implemented homogeneously and isothermally for the detection of enzymes, 30 DNAs, 31 RNAs, 32,33 and cancer cells 34 without the requirement of foreign enzymes, but they are rarely applied for the m 6 A assay. To the best of our knowledge, construction of a VMC10-mediated enzyme-free circuit for m 6 A detection has not been reported so far.…”