Integrated Colorisensing Platform with Microneedles and Metal-Phenol Nanozymes for Point-of-Care Testings of Acetylcholinesterase Activity and Its Drug Inhibitor.

Introduction: The detection of acetylcholinesterase (AChE) activity and the screening of its inhibitors are of significant importance for the diagnosis and drug therapy of nervous system diseases, particularly neurodegenerative disorders. This study aimed to develop a novel, integrated point-of-care testing (POCT) platform to address this need. Methods: We designed and integrated a colorimetric biosensor (Colorisensor) that combines a microneedle array with a metal-phenol nanozyme. The core sensing element is Iron (III)-polydopamine (Fe-PD) nanorods, which exhibit high peroxidase-like activity. The detection mechanism is based on the AChE-catalyzed hydrolysis of acetylthiocholine (ATCh) to produce thiocholine (TCh), which inhibits the nanozyme's activity. This inhibition prevents the catalytic oxidation of the chromogenic substrate TMB, leading to a measurable color change. A smartphone was utilized to quantify this change via red, green, and blue (RGB) values, creating a rapid and user-friendly platform for detections of AChE activity and its drug inhibitor. The nanorods and microneedle arrays were characterized using scanning electron microscopy, high-resolution transmission electron microscopy, energy-dispersive X-ray spectroscopy, ultraviolet-visible spectrophotometer, water absorption expansion rate, as well as mechanical property tests. Results and Discussion: The proposed Colorisensor demonstrated excellent analytical performance, including high selectivity and sensitivity with a low detection limit (LOD) of 0.007 mU/mL and a broad linear range from 0.01 to 1000 mU/mL. It was successfully applied to screen berberine hydrochloride as an AChE inhibitor. Crucially, the Colorisensor showed comparable accuracy to the standard Ellman's method and outperformed both traditional assays and emerging nanomaterial-based colorimetric methods by offering a wider detection range and a lower LOD. Conclusion: This study presents a successful proof-of-concept for an integrated microneedle and nanozyme-based Colorisensor. The platform provides a viable and promising alternative pathway for the early diagnosis of neurodegenerative diseases and the screening of therapeutic drugs, highlighting its significant potential for point-of-care applications.