LanthaScreen® Time-Resolved Fluorescence Energy Transfer Farnesoid X Receptor Coactivator Assay

The LanthaScreen® TR-FRET FXR Coactivator Assay (goat, PV4833, ThermoFisher Scientific) was performed to assess the affinity of tested compounds to FXR ligand-binding domain (LBD) in agonistic and antagonistic models. In the antagonistic assays, the FXR LBD was co-incubated with OCA or GW4064, together with tested ligands in increasing concentrations. The assay was performed according to the manufacturer’s protocols. After an 1 h incubation period at room temperature, the TR-FRET ratio of 520/495 nm was measured using the Biotek plate reader and used to calculate the IC₅₀ values from the concentration-response curves of each compound using GraphPad Prism version 9.1.0. software. Data have been obtained from three independent experiments performed in 4 replicates.