CRISPR/Cas12a-activated nanozyme-Aptasensor for ultrasensitive colorimetric detection of ochratoxin a based on MOF@pt@MOF nanostructures

Ochratoxin A (OTA), a prevalent cereal mycotoxin, exhibits carcinogenic and teratogenic properties that threaten public health. Consequently, sensitive and reliable OTA detection methods are critically needed. A CRISPR-Cas12a-mediated colorimetric biosensor was developed, integrating Cas12a regulation via modular hairpin structures with MOF@Pt@MOF nanozyme amplification. Hairpin structures incorporate target-specific sequences; target binding induces rearrangement, preventing Cas12a recognition while enabling dsDNA activators to restore Cas12a function. Magnetic beads (MBs) were coupled with MOF@Pt@MOF nanozymes via ssDNA to construct MBs-MOF@Pt@MOF probes. Following OTA-induced activation, Cas12a cleaves ssDNA, releasing nanozymes into the supernatant, catalyzing TMB oxidation to generate a blue chromophore. A detection limit of 0.055 ng/mL was achieved with excellent selectivity. This approach utilizes inhibitory aptamer conformational changes for precise Cas12a regulation and nanozyme signal amplification to enhance sensitivity and specificity. The modular design demonstrates versatility for diverse target detection in point-of-care testing (POCT) applications.