Journal
ANALYTICAL CHEMISTRY
Volume 92, Issue 12, Pages 8614-8622Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acs.analchem.0c01949
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Funding
- National Key Research and Development Program of China [2016YFA0201301]
- National Natural Science Foundation of China [21627808, 21721003]
- Key Research Program of Frontier Sciences, CAS [QYZDY-SSW-SLH019]
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In this study, we proposed a ratiometric electrochemiluminescent (ECL)/electrochemical (EC) biosensor based on duplex-specific nuclease (DSN)-assisted target recycling and multilayer catalytic hairpin assembly (CHA) amplification cascades for the detection of microRNA (miRNA). The DSN-assisted target recycling transformed miRNAs into a large number of ssDNA, which then catalyzed a multilayer CHA amplification cascade to produce numerous long dsDNA duplexes H-n/Hn+1 (n = 2, 4, 6, ...). Then the H-n/Hn+1 displaced the ferrocene (Fc)-labeled ssDNA (Sx+1, x = 1, 3, 5, ...) to hybridize with the S-x sequence on the gold electrode surface. Consequently, a great number of long S-x/H-n/Hn+1 duplexes were immobilized for binding Ru(phen)(3)(2+) to obtain an amplified ECL signal. Meanwhile, the EC signal of Fc was reduced, and the quenching effect of Fc to ECL signal also decreased. By measuring the ratio of the ECL signal of Ru(phen)(3)(2-) to the EC signal of Fc, quantitative analysis of miRNA-499 with high accuracy and reproducibility was obtained. The ratiometric biosensor shows high sensitivity and a wide linear range of 6 orders of magnitude. With the help of DSN-assisted target recycling, this strategy can be easily extended to detect other miRNAs without redesigning the CHA cascade system. The proposed hybrid ratiometric ECL/EC strategy enriches the ratiometric sensors and can find extensive applications in bioanalysis, especially for multiplex detection.
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