radioligandbindingsassays

Radioligand binding assays are biochemical techniques used to study the interaction between a radiolabeled ligand and its binding site, typically a receptor or transporter, in membranes or intact cells. The most common radioligands are radioactive isotopes such as tritium (3H) and iodine-125 (125I). The assay measures specific binding by separating receptor-bound radioligand from free radioligand, usually by filtration or precipitation, followed by quantification of radioactivity with a scintillation counter.

In a typical saturation binding experiment, receptor-rich preparations are incubated with increasing concentrations of radioligand to generate a saturation curve. Non-specific binding is determined in the presence of an excess unlabeled ligand and is subtracted from total binding to yield specific binding. Data analysis provides Bmax, a measure of receptor density, and Kd, the equilibrium dissociation constant reflecting affinity. Competition binding assays fix the radioligand concentration and vary concentrations of unlabeled competitors to estimate the affinity of other ligands. Kinetic binding assays measure association and dissociation rates to derive on- and off-rates.

Applications include pharmacological characterization of receptors, determination of receptor density in tissues, ligand selectivity profiling, and early-stage drug discovery. Radioligand binding can quantify high-affinity interactions with high sensitivity and requires relatively small sample amounts.

Limitations include reliance on radioactive materials, potential artifacts from receptor preparation, and non-specific binding. Safety and regulatory considerations demand appropriate licensing, training, shielding, and proper waste disposal. Despite these constraints, radioligand binding assays remain a foundational tool for quantitative receptor pharmacology.