Construction of High-Performance Wash-Free Aβ Probes Based on Amino Group Optimization and Amphiphilic Molecular Engineering Strategies

β-Amyloid (Aβ) protein is a misfolded protein that plays a pivotal role in the pathogenesis of Alzheimer's disease (AD) and is recognized as one of the primary pathological hallmarks of this disorder. Despite significant advancements in developing fluorescent probes for Aβ detection, research on the specific impact of end-group substituents on probe performance and the design of high-performance, wash-free Aβ probes remains limited. In this study, we designed and synthesized a series of water-soluble Aβ fluorescent probes, X-PYOH and X-PYC6, using a styrene pyridinium cation as the core structure by varying the end-group substituents. We systematically investigated their hydrophilicity/hydrophobicity, optical properties, Aβ detection capability, and imaging performance. The results demonstrated that among the various amino-substituted X-PYOH probes, PL-PYOH exhibited superior Aβ sensing, affinity, anti-interference capacity, Aβ plaque labeling performance, and wash-free characteristics. In contrast, PZ-PYOH and CT-PYOH showed poor Aβ detection performance. By modification of the substituents at the pyridinium cation end to decrease their hydrophilicity and preparation of X-PYC6 (PZ-PYC6, CT-PYC6), the responsiveness and affinity to Aβ fibrils were significantly enhanced, and the labeling performance of Aβ plaques was markedly improved. In conclusion, optimizing the types of amino substituents and balancing hydrophilicity and hydrophobicity can lead to the development of high-performance, wash-free imaging Aβ probes.