Thioflavin T assay for Aβ fibril formation

First, a 2.5 mM ThT stock solution was prepared by adding 8 mg of ThT to 10 ml of 1× PBS buffer and filtering through a 0.2 μm syringe filter. This solution was stored in the dark. This stock solution was diluted into the 1× PBS buffer on the day of analysis to generate a 20 μM ThT working solution and covered with aluminum foil. Then, 1 mg of Aβ 25–35 (Sigma Aldrich, Korea) was dissolved in DMSO to the final concentration of 10 mM. This Aβ stock solution was diluted into 1× PBS buffer to generate 10, 50, and 100 μM Aβ solutions and incubated at RT with gentle shaking for an appropriate amount of time. On the day of analysis, 5 μl of the ThT working solution was mixed with 95 μl of each Aβ fibril solution in a black 96-well plate and measured using a microplate reader (Victor3, Perkin Elmer, USA). The fluorescence intensity was measured by excitation at 450 nm and emission at 485 nm.