Signal-off Electrochemiluminescence Biosensor Based on Phi29 DNA Polymerase Mediated Strand Displacement Amplification for MicroRNA Detection

Abstract

A target induced cycling strand displacement amplification (SDA) mediated by phi29 DNA polymerase (phi29) was first investigated and applied in a signal-off electrochemiluminescence (ECL) biosensor for microRNA (miRNA) detection. Herein, the target miRNA triggered the phi29-mediated SDA which could produce amounts of single-stranded DNA (assistant probe) with accurate and comprehensive nucleotide sequence. Then, the assistant probe hybridized with the capture probe and the ferrocene-labeled probe (Fc-probe) to form a ternary "Y" structure for ECL signal quenching by ferrocene. Therefore, the ECL intensity would decrease with increasing concentration of the target miRNA, and the sensitivity of biosensor would be promoted on account of the efficient signal amplification of the target induced cycling reaction. Besides, a self-enhanced Ru(II) ECL system was designed to obtain a stable and strong initial signal to further improve the sensitivity. The ECL assay for miRNA-21 detection is developed with excellent sensitivity of a concentration variation from 10 aM to 1.0 pM and limit of detection down to 3.3 aM.

Emerging Biosensing Approaches for microRNA Analysis

“…Enzymatic amplification strategies have been deployed using rolling circle amplification to form DNAzymes, 78 cyclic exponential amplification recycling, 79 doxorubicin conjugated quantum dots, 80 T7 exonuclease recycling and downstream silver deposition, 81 ECL quenching via Phi29 DNA polymerase mediated strand displacement, 82 and a dual target amplification strategy with combined ECL and fluorescence detection. 83 With one exception, all of these strategies report LODs ranging between 10 fM and 100s of aM with dyamic ranges varying 2–5 orders of magnitude.…”

Highly Efficient Intramolecular Electrochemiluminescence Energy Transfer for Ultrasensitive Bioanalysis of Aflatoxin M1

“…The conventional three-electrode system was used as in our previous work. [31] Cyclic voltammetric (CV) and electrochemical impedance spectroscopy (EIS) were carried out by aC HI 610A electrochemistry workstation (Shanghai CH Instruments, China). The surface morphology of PtNPs were characterized by scanning electron microscopy (SEM, S-4800, Hitachi, To kyo, Japan).…”

A Novel Surface‐tethered Analysis Method for Mercury (II) ion Detection via Self‐assembly of Individual Electrochemiluminescence Signal Units

“…The synthesis procedure of GO referred to a modified Hummers method , and GO@PTCA‐TAEA we prepared by the previous description with some modifications . First, an amount of 5 mg PTCDA was dissolved in 40 mL 0.01 M sodium hydroxide solution, and then, the color of the solution was turned to pale yellow with slight green fluorescence.…”