A specially designed DNA-assembled framework structure probe coupled with loop-mediated isothermal amplification (LAMP)-DNA signal transducer for rapid and sensitive electrochemical detection of miRNA

Herein, a new DNA-assembled framework structure (DAFS) probe coupled with LAMP-DNA signal transducer is developed for rapid and sensitive detection of miRNA let-7a. The DAFS probe was formed via self-assembly of capture strand (P) and ferrocene-labelled DNA strands (P1, P2) and was immobilized via hybridization of P1 and P2 with the DNA strands tethered to electrode surfaces. On the other hand, forward primer (FP) of LAMP was hybridized with assistant strand (AS) attached on Au@Fe 3 O 4 composite nanoparticles. After LAMP reaction started, the duplex FP:AS was separated to expose the AS. Then, with a facile magnetic separation, the exposed AS was collected and further cut by Mn 2+ , outputting the signal transducer T*. The DAFS probe could disassemble by the T* in the presence of two hairpin DNA probes (H1, H2) and release the P1 and P2 into the solution, causing an electrochemical signal change for realizing sensitive detection of final target let-7a. The electrochemical detection could complete within 50 min with a good linear relationship from 100 aM to 20 pM and a detection limit of 48 aM. Our biosensors performed well in human serum spiked with the let-7a and the let-7a extracts obtained from the human breast cancer cell (MCF-7). • The LAMP-DNA signal transducer was collected by using Au@Fe 3 O 4 NPs for electrochemical detection, minimizing possible interferences from the complex components in the LAMP reaction solution containing real bio-samples. • The capture strand of the DAFS probe was located at the outmost side of the surface-tethered DAFS probe modification layer, therefore its disassembly to release the Fc-labelled strands was fast, and the response to the target was rapid. • This study demonstrates the promising potential of the DAFS probe in designing electrochemical biosensors for rapid detection of nucleic acids.