Photophysical study of Thioflavin T as fluorescence marker of amyloid fibrils

•A detailed photophysical analysis of Thioflavin T in various media is presented.•The strong wavelength dependency and enhancement of the fluorescence is explained.•Solvent dependent ground-state conformations determine the fluorescence properties.•Affinity and specificity to amyloid-β (1–42) fibrils and BSA are compared.•The use of Thioflavin T for quantitative determinations of Aβ42 fibrils is assessed.

Thioflavin T is a highly sensitive fluorescent marker of amyloid fibrils that has been widely used for in vitro biomedical assays. However, neither its complex photophysical behavior nor its binding mode to amyloid fibrils are still well understood. We present a detailed analysis of the photophysical properties of Thioflavin T in various media, including solvents and solvent mixtures of different viscosities as well as fibrillar and globular proteins. We propose a model that explains the strong wavelength dependency of the Thioflavin T fluorescence and the large fluorescence enhancement in certain environments. We determine the binding affinities and the fluorescence properties of Thioflavin T bound to amyloid-β (1–42) fibrils and to bovine serum albumin and discuss the sensitivity and the specificity of this probe to amyloid aggregates. These results allow us to assess the suitability of Thioflavin T for quantitative determinations in biomedical studies.

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