3D DNA nanonet structure coupled with target-catalyzed hairpin assembly for dual-signal synergistically amplified electrochemical sensing of circulating microRNA

DNA nanomaterials are reliable and powerful tools in the development of a variety of biosensors owing to their notable self-assembly ability and precise recognition capability. Here, we propose a DNA nanomaterial-based system for the dual-amplified electrochemical sensing of circulating microRNAs by a coupled cascade of catalyzed hairpin assembly (CHA) and three-dimensional (3D) DNA nanonet structure. In the target-assisted CHA process, the stable hairpin structures H1 and H2 act as probes for the recognition and recycling of circulating microRNAs, leading to the formation of abundant H1–H2 duplexes with tails. Subsequently, a 3D DNA nanonet structure was introduced, which was assembled using three DNA strands constructed X-DNA monomers as the building blocks, and hybridized to the tails of H1–H2 duplexes. The successful integration of target-assisted CHA and 3D DNA nanonet structure induced the second signal amplification. The designed biosensor performed under optimized experimental conditions, and exposed admirable analytical performance for the detection of circulating miR-21, with a wide linear range from 10 fM to 1 nM, high sensitivity of limit of detection (LOD) of 3.6083 fM, good specificity in the face of single nucleotides and other microRNAs, satisfactory stability and reproducibility for practical analysis. Furthermore, the clinical applicability for circulating miR-21 detection was verified in human serum samples without additional treatment. We hope that this elaborated biosensor will provide new opportunities for bioassays based on DNA nanomaterials.