Multiple Amplified Electrochemical Detection of MicroRNA‐21 Using Hierarchical Flower‐like Gold Nanostructures Combined with Gold‐enriched Hybridization Chain Reaction

Abstract

In this work, we present a multiple amplified electrochemical method that can detect target microRNA‐21 (miRNA‐21) with high sensitivity by using hierarchical flower‐like gold nanostructures (HFGNs) and gold‐enriched enzyme‐free amplification. The HFGNs deposited on ITO conducting glass served as the electrode substrate with large specific surface area, which allowed the immobilization of large amounts of capture DNA (DNA‐1). Then, the gold‐enriched hybridization chain reaction (HCR) for signal amplification was attached onto the electrode through target miRNA‐mediated sandwich hybridization. The initial strands (I‐DNA) enriched on gold nanoparticles (AuNPs) provided plenty of sites for the triggering of the HCR reaction after the addition of H1 and H2 strands, which provided exponential increase of binding sites for the loading of electrochemical signal molecules [Ru(NH3)6]3+. Therefore, multiple signal amplification could be achieved to reach an ultrasensitive detection of miRNA‐21. A wide linear dynamic range from 1 fM to 1 nM and a detection limit of 0.12 fM (S/N=3) was reached with good selectivity. The electrochemical assay method in this work may hold a great potential for clinical diagnosis of genetic disease in the future.

Intrinsic Electrocatalysis of RNA as a Label‐free and Reagent‐less Tool for Detection of MicroRNAs

“…Redox SW peak P enabled us to measure RNA down to 1 nM concentration (Figure D 1 ), making it less sensitive than catalytic CPS peak H. CPS is thus better suited for sensitive measurements than SWVS or CV, because catalytic peaks obtained with CPS at low current densities arise from the exchange of many electrons per RNA molecule, as compared to 2–4 electron exchange during redox processes (using peak CA or peak P). Of course, sensitivity of our label‐free reagent‐less approach cannot compete with various label‐based assays for miRNA detection, often reaching LODs in femtomolar or even attomolar range . On the other hand, we have recently introduced electrochemical bioplatform for miR‐21 detection in clinical samples (we used samples from women suffering from cervical precancerous lesions), which was based on specific S9.6 antibody capturing miRNA‐DNA duplexes and hybridization chain reaction .…”

Nanotechnology in emerging liquid biopsy applications

“…The detection was finally accomplished through the electroactive compound Ru(NH3) 6 3+ . The detection limit of the method was 0.12 fM of synthetic miRNA target [ 26 ]. Alternatively, in another approach, target miRNA triggered a target chain displacement polymerization reaction by DNA Klenow fragment, while the electrochemiluminescence signal was generated by a DNA/Ru(bpy) 3 2+ /AuNPs complex, leading to a great signal enhancement with an LOD of 43 aM [ 46 ].…”

“…Nanomaterials have been significantly valuable in liquid biopsy applications for signal enhancement, as they do not require use of enzymatic reactions or multiple enhancement steps. Signal enhancement is mainly attributed to their high surface-to-volume ratio and their excellent optical and electrical properties [ 26 ]. The typical approach to improve the detectability of a sensor is to increase the surface-to-volume ratio along with the improved signal activity [ 33 ].…”

Nanomaterials for molecular signal amplification in electrochemical nucleic acid biosensing: recent advances and future prospects for point-of-care diagnostics

“…In this manner, a 32-fold enhancement in detection limits has been reported when compared to a single gold nanoparticle label. 120,121 On the other hand, a unique approach for miRNA detection was recently described by Gooding and co-workers, as illustrated in Fig. 8.…”