BACE1 Activity Assay Protocol

Also, a β-secretase (BACE1) activity assay kit was purchased from Sigma-Aldrich (catalog number CS0010) and was used as described previously [26 (link)]. In this experiment, an enhancement of the fluorescence signal after substrate cleavage via BACE1 was observed [27 (link)]. The concentrations of compounds were the same as in the assay with AChE. The assay was performed using a black 96-well microplate. Fluorescence was measured on an Infinite M200 fluorescence spectrophotometer (TECAN, Männedorf, Switzerland) (ex. 320 nm; em. 405 nm) in triplicate experiments with a negative control (no enzyme) and a positive control (enzyme activity supplied). Secretase activity correlated with fluorescence value. Its 50% decrease was calculated as IC₅₀ (µM).