Allosteric interactions involve the simultaneous binding of two ligands to the same receptor. An allosteric modulator causes a conformational change in the receptor protein that yields a change in the binding or signaling of an orthosteric agent, i.e., an agonist or competitive antagonist that binds to the endogenous agonist binding site. Because of the complex nature of allosteric phenomena, the detection and quantification of their effects on orthosteric ligand binding relies on the use of both equilibrium and non-equilibrium assays to ensure proper interpretation of the findings. Outlined in this unit are the most common experimental approaches for measuring allosteric effects on orthosteric ligand affinity at G protein-coupled receptors. There is also a discussion of the analysis of experimental data derived from such assays. Curr. Protoc. Pharmacol. 52:1.22.1-1.22.41.