Description :
The TR-FRET Retinoid X Receptor alpha Coactivator Assay provides a sensitive and robust method for high-throughput screening of potential RXR alpha ligands as agonists of ligand-dependent coactivator recruitment. The kit uses a terbium (Tb)-labeled anti- GST antibody, a fluorescein-labeled coactivator peptide, and a human recombinant RXR alpha ligand-binding domain (RXR alpha-LBD) that is tagged with glutathione-S-transferase (GST) in a homogeneous mix-and-read assay format.
To assay:
When running the TR-FRET Retinoid X Receptor alpha Coactivator Assay in agonist mode (to identify agonist compounds), RXR alpha-LBD is added to ligand test compounds followed by addition of a mixture of the fluorescein-coactivator peptide and Tb-labeled anti-GST antibody. After an incubation period at room temperature, the TR-FRET ratio of 520/495 is calculated and can be used to determine the EC50 from a dose-response curve of the compound. Based on the biology of the RXR alpha-coactivator peptide interaction, this ligand EC50 is a composite value representing the amount of ligand required to bind to receptor, effect a conformational change, and recruit coactivator peptide.
Storage :
TR-FRET RXR alpha Coactivator Assay Kits contain RXR alpha-LBD (GST) protein, fluorescently labeled PGC1a coactivator peptide, Tb-labeled anti-GST antibody and buffers. Store components as indicated in the assay protocol (-80°C, -20°C or 4°C).
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