Low Background Cascade Signal Amplification Electrochemical Sensing Platform for Tumor-Related mRNA Quantification by Target-Activated Hybridization Chain Reaction and Electroactive Cargo Release

Abstract

Herein a low background cascade signal amplification electrochemical sensing platform has been proposed for the ultrasensitive detection of mRNA (mRNA) by coupling the target-activated hybridization chain reaction and electroactive cargo release from mesoporous silica nanocontainers (MSNs). In this sensing platform, the 5′-phosphate-terminated DNA (5′-PO 4 cDNA) complement to target mRNA is hybridized with the trigger DNA and anchor DNA on the surface of the MSNs, aiming at forming a double-stranded DNA gate molecule and sealing the methylene blue (MB) in the inner pores of the MSNs. In the presence of target mRNA, the 5′-PO 4 cDNA is displaced from the MSNs and competitively hybridizes with mRNA, which led to the liberation of the trigger DNA and the opening of the MSNs pore. The liberated trigger DNA can be then immobilized onto the electrode surface through hybridization with the capture DNA, triggering HCR on the electrode surface. At the same time, the MB released from the MSNs will selectively intercalate into the HCR long dsDNA polymers, giving rise to significant electrochemical response. In addition, due to the λ-exonuclease (λ-Exo) cleavage reaction-assisted target recycling, more amounts of trigger DNA will be liberated and trigger HCR, and numerous MB are uncapped and intercalate into the HCR products. As proof of concept, thymidine kinase 1 (TK1) mRNA was used as a model target. Featured with amplification efficiency, label-free capability, and low background signal, the strategy could quantitatively detect TK1 mRNA down to 2.0 aM with a linear calibration range from 0.1 fM to 1 pM. We have also demonstrated the practical application of our proposed sensing platform for detecting TK1 mRNA in real samples, opening up new avenues for highly sensitive quantification of biomarkers in bioanalysis and clinical diagnosis.

Electron Acceptors Co-Regulated Self-Powered Photoelectrochemical Strategy and Its Application for Circulating Tumor Nucleic Acid Detection Coupled with Recombinase Polymerase Amplification

“…To further characterize the construction process of PEC assay step by step, EIS was performed in a 5.0 mM [Fe­(CN) 6 ] 3–/4– solution containing 0.1 M KCl. [Fe­(CN) 6 ] 3–/4– was used as a negatively charged redox label, and the semicircle diameter of impedance corresponded to the electron transfer resistance (Ret) . As shown in Figure B, the bare GE displayed a small Ret value (curve a).…”

Long noncoding RNAs: from genomic junk to rising stars in the early detection of cancer

“…The increase in the specific electrochemical signal by cascade amplification could also be accompanied by an increase in the background signal, albeit not necessarily to the same extent. To circumvent this issue, it has been recently proposed an electrochemical genosensing platform that combines an HCR amplification on gold substrates modified with a binary DNA self-assembled monolayer, with target-responsive electroactive cargo (methylene blue, MB) release from mesoporous silica nanoparticles (MSNs) [36]. Methylene blue molecules, chosen to reveal the HCR by intercalation into the long ds-DNA molecules generated on the gold…”

Mesoporous Silica Containers and Programmed Catalytic Hairpin Assembly/Hybridization Chain Reaction Based Electrochemical Sensing Platform for MicroRNA Ultrasensitive Detection with Low Background

“…Very recently, we presented an MSNs-based sensing platform for the ultrasensitive detection of nucleic acid biomarker by involving the exonuclease-induced target recycling amplification . The exonucleases were highly efficient to power the nucleic acids amplification.…”

“…Very recently, we presented an MSNs-based sensing platform for the ultrasensitive detection of nucleic acid biomarker by involving the exonuclease-induced target recycling amplification. 30 The exonucleases were highly efficient to power the nucleic acids amplification. However, such a nucleases-based amplification strategy might suffer from the inhibiting factors in complex samples analysis, such as buffers, temperature, and other inhibitors.…”