Dysregulation of microRNA (miRNA) expression is associated with a variety of human diseases, including cancers, and the development of highly sensitive miRNA detection is important for the early diagnosis of cancer. In this study, we developed an electrochemical biosensing platform for miRNA detection through the integration of a DNA-gated metal-organic framework (MOF) signal probe, duplex-specific nuclease (DSN)-assisted signal amplification, and a CRISPR/Cas12a system. The zirconium-based MOF UiO-66-NHwas engineered as a nanocarrier for methylene blue (MB) entrapment, and single-stranded DNA (ssDNA) was capped on the MOF surface as a "gatekeeper" that allowed the controlled release of MB molecules, resulting in a DNA-gated MOF electrochemical signal probe. In the presence of target miRNA, target-initiated DSN cyclic amplification and digestion of the CRISPR/Cas12a initiator DNA sequence blocked the collateral cleavage activity of Cas12a toward ssDNA, thus preventing the release of MB and generating a high electrochemical signal. Conversely, the CRISPR/Cas12a system would be activated to cleave ssDNA in the absence of the target, leading to the release of a signal molecule and a low response. With the usage of the proposed biosensing strategy, sensitive detection of miRNA let-7a, a biomarker associated with nonsmall cell lung cancer, has been achieved. Therefore, this work expands the application scope of a MOF as a nanocarrier to prepare electrochemical signal probes and provides a valuable biosensing method for clinical diagnosis. 2
