Incorporating CRISPR Techniques with Electrochemical Sensors: From Signal-Off to Signal-On Assays

Electrochemical biosensors integrating the programmable nucleic acid recognition of CRISPR systems with the low-cost and portable electrochemical transduction have emerged as powerful sensing platforms for molecular diagnostics. Early designs mainly relied on turn-off signal transduction, where target-activated CRISPR enzymes cleave probes conjugated to the electrode, resulting in the release of redox reporters from the electrode surface and the consequent signal decrease. Although conceptually straightforward, these turn-off sensors are intrinsically limited by high background, low sensitivity, and large signal variations. To address these limitations, recent efforts have increasingly shifted toward turn-on strategies, in which electrochemical signals are generated in response to target binding. This review highlights recent advances in applying CRISPR technology to electrochemical biosensing, with a focus on the design principles of CRISPR systems and molecular assembly to achieve turn-off and turn-on signal transduction. Nanomaterials, DNA nanotechnology, and amplification strategies facilitate emerging turn-on approaches for sensitive electrochemical sensing. Key challenges and research needs include improving the limit of detection, robustness, and applicability to point-of-care and on-site testing. This review emphasizes the importance of signal-transduction designs and provides perspectives for developing sensitive, specific, and practical CRISPR-based electrochemical biosensors.