Advancing the Validation of the Enrichment-Enhanced Detection Strategy with Au Nanoclusters for AChE Detection.

High-sensitivity fluorescent probes provide a powerful tool for understanding life processes and functioning mechanisms. Therefore, the development of a universal strategy to optimize probes holds substantial importance. Herein, we developed a novel strategy for common probe upgrades: rather than simply pursuing a higher fluorescence intensity of the probe itself, we tried to promote the detection sensitivity by enhancing the probe-substrate interactions. Fortified with polyionic polymers, self-assembled probes could be endowed with enhanced attractions to the substrate. In this work, we took the AChE-AuNCs detection system as a typical and important example to verify this concept of the "enrichment-enhanced detection" strategy (EED strategy). Two probes, AuNCs@GC and AuNCs@CMCS, with similar composing polymers (chitosan derivatives), microstructures, fluorescence profiles, and distinct charges were delicately designed and thoroughly studied. CMCS with an abundance of negatively charged carboxy groups plays an important role in the enrichment of thiocholine through electrostatic interactions. Thus, despite having similar composing components, structures, and almost identical fluorescence profiles, the negatively charged composite shows superior sensitivity (15.2-fold enhancement) and response time (2-fold faster) compared to the AuNCs@GC, thereby validating the feasibility of the EED strategy. Overall, our work validates the EED strategy and applies it to the accurate detection of AChE activity. We believe that this strategy offers substantial insights for the generalization and enhancement of advanced nanoprobes.