Tailoring high-energy self-powered sensing system by Walker-mediated CRISPR/Cas12a cascade signal amplification and hybridization chain reaction for ultrasensitive microRNA detection

MicroRNAs (miRNAs) are important cancer-associated biomarkers for early detection. However, accurately measuring miRNAs is challenging due to their low abundance and high homogeneity in human serum. CRISPR technology can greatly improve the specificity and sensitivity of detection in biosensor construction. In this study, we develop a remarkably responsive self-powered biosensing platform for detecting microRNA-21 (miRNA-21) using hybridization chain reaction (HCR) and CRISPR/Cas12a amplification cascade mediated by 3D DNA walkers. Carbon-coated molybdenum disulfide nanotubes with a large surface area and excellent conductivity are fabricated on flexible carbon paper as the anode and cathode for biological fuel cell construction. Glucose oxidase is loaded onto the anode surface, while serving as the biological cathode, it initiated HCR, generating abundant double-stranded DNA. The negatively charged DNA backbone captures electron acceptors [Ru(NH3)6]3+ through electrostatic adsorption, resulting in a significant open circuit voltage (EOCV) signal. Through CRISPR/Cas12a amplification cascade mediated by 3D DNA walkers, a smaller EOCV is produced, enabling sensitive target detection. The method exhibits a detection range spanning from 0.0001 to 10,000 pM and capable of detecting as low as 19.0 aM (S/N = 3). This highly selective and sensitive method provides an effective approach for accurately detecting tumor markers in clinical applications.