A competitive binding assay typically measures the binding of a labeled ligand to a target protein in the presence of a second, competing but unlabeled ligand. This assay can be used to assess qualitative binding information as well as relative affinities of two or more molecules for one target. It can also be used to quantify high-affinity interactions that are difficult to measure directly.
The main requirement for this assay is that the given molecules bind to the same site of the target protein. The unlabeled competitor then competes for binding to the same site with the labeled molecule. This assay yields the EC50, the concentration that displaces 50% of the labeled molecule from the binding site. The affinity of the non-labeled ligand for the protein can then be calculated. Reach out to the NanoTemper Technologies Support Team (NanoTemper Technologies Support) for help with setting up competitive binding assays.