Solvatochromic\nFluorescent Probe for Visualizing Protein\nAggregation via STED Imaging

Elucidating the structure of protein aggregates is vital\nfor overcoming\nhuman diseases arising from protein misfolding. It is rarely reported\nthat the conventional protein-amyloid fibrillation probe ThT can be\nused as a STED agent to visualize protein aggregation structures at\nthe nanometer level, suffering from small Stokes’ shift and\nphotobleaching. Herein, we report a donor–acceptor (D–A)-type\nfluorophore, IAD2, with red emission and a large Stokes' shift\n(196\nnm). It is demonstrated that IAD2 exhibited a 7.8-fold enhancement\non the fluorescence intensity and an obvious 94 nm blue shift on the\nemission wavelength after binding with bovine insulin fibril, due\nto the alternation of the microenvironment. Besides, IAD2 has a relatively\nhigher binding affinity to insulin fibrils in comparison to ThT. Molecular\ndocking assay also verified the binding sites and interaction forces\nbetween IAD2 and insulin fibrils. Owing to its high depletion efficiency\nand good photostability, IAD2 was utilized to stain insulin fibrils\nto achieve STED imaging with a resolution of 101.2 nm. This study\nwill shed light on the design of novel solvatochromic fluorescent\nprobe for the super-resolution imaging of protein aggregate structure.