A Solvatochromic Fluorescent Probe Reveals Polarity Heterogeneity upon Protein Aggregation in Cells

Abstract We report a crystallization‐induced emission fluorophore to quantitatively interrogate the polarity of aggregated proteins. This solvatochromic probe, namely “AggRetina” probe, inherently binds to aggregated proteins and exhibits both a polarity‐dependent fluorescence emission wavelength shift and a viscosity‐dependent fluorescence intensity increase. Regulation of its polarity sensitivity was achieved by extending the conjugation length. Different proteins bear diverse polarity upon aggregation, leading to different resistance to proteolysis. Polarity primarily decreases during protein misfolding but viscosity mainly increases upon the formation of insoluble aggregates. We quantified the polarity of aggregated protein‐of‐interest in live cells via HaloTag bioorthogonal labeling, revealing polarity heterogeneity within cellular aggregates. The enriched micro‐environment details inside misfolded and aggregated proteins may correlate to their bio‐chemical properties and pathogenicity.