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A Universal Ratiometric Method for Micro-Rna Detection Based on the Ratio of Electrochemical/Electrochemiluminescence Signal, and Toehold-Mediated Strand Displacement Amplification Publisher Pubmed



Niazi M1 ; Azizi A1 ; Khajavi Z1 ; Sheikh M1 ; Taheri S2 ; Radfar S2 ; Alizadeh A3 ; Ghanbari R1
Authors
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Authors Affiliations
  1. 1. Department of Biological Science and Technology, Najafabad Branch, Islamic Azad University, Najafabad, Iran
  2. 2. Stem Cell and Regenerative Medicine Center of Excellence, Tehran University of Medical Science, Tehran, Iran
  3. 3. Department of Organic Chemistry, Faculty of Chemistry, Alzahra University, Tehran, 1993893973, Iran

Source: Analytica Chimica Acta Published:2023


Abstract

An ultra-selective and reproductive ratiometric platform was introduced based on the ratio of Ru(phen)32+ electrochemiluminescence (ECL) signal and methylene blue (MB) electrochemistry (EC) signal, which was amplified using a specific and efficient toehold-mediated strand displacement (TMSD). The stable DNA nanoclews (NCs) were efficiently loaded with MB (MB-NCs) as EC signal tags after being synthesized utilizing a simple rolling circle amplification reaction. Besides, Ti3C2-based nanocomposite could apply as a superb carrier for both Ru(phen)32+ and gold nanoparticles (Ti3C2–Au–Ru), resulting in a nearly constant ECL internal reference to eliminate the possible interferences. The Ti3C2–Au–Ru was attached to the surface of the electrode using Nafion, which exhibited excellent conductivity, and hairpin DNAs (hDNAs) were fixed on AuNPs via an Au–S bond. The designed biosensor was finally applied for miRNA-18a detection as a target model. The TMSD method made it possible to concurrently convert and amplify a single miRNA-18ainput into a large amount of output DNAs with high selectivity. These output DNAs were designed to unfold the stem-locked area of hDNAs. The opened hDNAs then hybridized with the MB-NCs to produce an EC signal. In the proposed biosensing system, by raising the target concentration of miRNA, the EC signal gradually rose, the ECL signal remained nearly constant, and the ratiometric detection method markedly promoted biosensor accuracy. Linear correlations of the ratio value of the EC/ECL with miRNA-18a concentrations between 20 aM and 50 pMwere observed, with the limit of detection of 9 aM. The biosensor was applied to detect miRNA-18a in real serum samples with satisfactory results. © 2023 Elsevier B.V.
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