CRISPR/Cas13a Powered Portable Electrochemiluminescence Chip for Ultrasensitive and Specific MiRNA Detection

MicroRNAs (miRNAs) have been widely investigated as potential biomarkers for early clinical diagnosis of cancer. Developing an miRNA detection platform with high specificity, sensitivity, and exploitability is always necessary. Electrochemiluminescence (ECL) is an electrogenerated chemiluminescence technology that greatly decreases background noise and improves detection sensitivity. The development of a paper-based ECL biosensor further makes ECL suitable for point-of-care detection. Recently, clustered regularly interspaced short palindromic repeats (CRISPR)/Cas13a as high-fidelity, efficient, and programmable CRISPR RNA (crRNA) guided RNase has brought a next-generation biosensing technology. However, existing CRISPR/Cas13a based detection often faces a trade-off between sensitivity and specificity. In this research, a CRISPR/Cas13a powered portable ECL chip (PECL-CRISPR) is constructed. Wherein target miRNA activates Cas13a to cleave a well-designed preprimer, and triggers the subsequent exponential amplification and ECL detection. Under optimized conditions, a limit-of-detection of 1 × 10^-15 m for miR-17 is achieved. Through rationally designing the crRNA, the platform can provide single nucleotide resolution to dramatically distinguish miRNA target from its highly homologous family members. Moreover, the introduction of "light-switch" molecule [Ru(phen)₂dppz]²⁺ allows the platform to avoid tedious electrode modification and washing processes, thereby simplifying the experimental procedure and lower testing cost. Analysis results of miRNA from tumor cells also demonstrate the PECL-CRISPR platform holds a promising potential for molecular diagnosis.