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Assay Method Information

Assay Name:  Radioligand Binding Assay
Description:  Compounds of the invention were assessed in a competition binding assay where different concentrations of compounds were incubated with the LXR ligand binding domain (LBD) in the presence of radiolabeled LXR ligand [3H]TO901317. The amount of the LXR-LBD that complexed with [3H]T0901317 was measured by scintillation proximity assay (SPA) employing non-specific binding of LXR-LBD to poly-lysine coated Yttrium silicate beads. Partially purified LXR alpha or beta LBD protein (15-45 nM) was incubated at rt for 30 min with 15 nM [3H]T0901317 (25-40 Ci/mmol) and different concentrations of test compounds in 80 uL of phosphate buffered saline (PBS) buffer containing 2.5% DMSO, 1% glycerol, 2 mM EDTA, 2 mM CHAPS and 5 mM DTT in 96-well plates. Poly-lysine SPA beads (50 ug) were added to each well and the total volume was adjusted to 120 uL. The plates were shaken on an orbital shaker for 20 min and then allowed to settle for 10 more minutes at room temperature.
Affinity data for this assay
 

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Last update November 1, 2007
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