Synergy ht multi reader

The aggregation of Aβ₄₂ monomers was measured by Thioflavin T (ThT) assay. For dose-response experiments, 10 μM of rAβ₄₂ were combined with or without several concentrations (30, 10, 3, 1 μM) of compounds, in a 96 well black plate. Plates were incubated for 48 hours at 37 °C and then, ThT was added at a final concentration of 20 μM. The fluorescence was measured using a Synergy HT multi-reader from Biotek (Winooski, VT) at 450 nm for excitation and 485 nm for emission. Fluorescence intensity values were determined as the delta (Δ) of ThT fluorescence in the presence of amyloid by subtracting the basal fluorescence of ThT alone. The assay was performed in a final volume of 200 μL and samples were prepared in duplicates in the presence of DMSO (0.5 %). Morin (100 μM) was used as inhibition control.
For time-course experiments, the SensoLyte Thioflavin T β - Amyloid (1-42) aggregation kit was used, following the manufacturer instructions. Compounds were used at a concentration of 30 μM. The ThT fluorescence intensity of each reaction was measured from the top of the plate every 10 min using Synergy HT multi-reader from Biotek (Winooski, VT) with 450/485-nm excitation/emission filters set with 15 seconds of shaking at 100 rpm before each read. The fluorescence intensity values were determined as described above.