ECL and visual dual-mode detection of miRNA-21 based on HRP/Au NPs composite probe and CRISPR/Cas12a strategy

Abstract

Signal-amplification sensing strategies for rapid and accurate detection of tumor marker miRNA is of great practical significance for cancer prevention and treatment. Catalytic hairpin assembly (CHA) nucleic acid amplification reaction and CRISPR/Cas12a strategies have exhibited huge prospect due to their flexible design and efficient signal amplification. We constructed an electrochemiluminescence (ECL) biosensor based on horseradish peroxidase/gold nanoparticles (HRP/Au NPs) nanocomposite probe and CRISPR/Cas12a (LbCpf1), and completed a dual-mode detection of miRNA-21. When the target RNA is present in the system, the CHA) cycle was initiated, and the double-stranded DNA containing a specific protospacer adjacent motif (PAM) base sequence was generated to successfully activate the trans-cut activity of CRISPR/Cas12a. After the prepared HRP/Au NPs nanocomposite probe was cleaved by CRISPR/Cas12a, the supernatant containing HRP was collected through magnetic separation and subsequently introduced to the cathode of the bipolar electrode facilitating anodic ECL luminescence response. In addition, the supernatant was added to the TMB chromogenic solution for chromogenic reaction. Thus, ECL detection and visual detection of miRNA-21 were realized. Dual-mode detection improves the accuracy of the sensor, and the utilization of bipolar electrodes on indium tin oxide (ITO) coated glass could also provide some useful references for further development of point-of-care detecting devices for early cancer prevention and diagnosis.