Photophysical Properties of Thioflavin T. Does it form Excimers When Integrated into Amyloid Fibrils?

Abstract

Thioflavin T (ThT) is a proven tool to detect and study of the amyloid fibrils that frequently accompany the development of serious human diseases, including Alzheimer's, Parkinson's, prion, and more than 25 other diseases. Obviously, the success of ThT use for investigation of fibrils largely depends on the correct understanding of its photophysical properties and mechanism of interaction with amyloid fibrils. Using the specially elaborated approach for recoded fluorescence intensity correction on the primary and secondary inner filter effects and spectrofluorimeter with the horizontal slits we for the first time observed the fluorescence band with a maximum at 570 nm for ThT aqueous solutions with high concentrations and proved that this band can be attributed to dye excimers. We showed that ThT excimer fluorescence spectrum has nothing in common with the fluorescence of ThT incorporated into amyloid fibrils, which has maximum at about 490 nm. Furthermore we proved that the increase in the ThT fluorescence intensity when the dye binds to amyloid fibrils can be explained only by the molecular rotor nature of ThT. Thus, we concluded that there is no basis to suggest that the fluorescence of ThT bound to fibrils is caused by its excimers.This work was supported in part by grant 14-08-90024_Bel (K.K.T.), 13-04-02068 (A.I.S.) from the Russian Foundation of Basic Research, the Program “Molecular and Cell Biology” of the Russian Academy of Sciences, and RF President fellowship SP-1982.2015.4 (A.I.S.).