Radioligandbinding

Radioligand binding is a biochemical technique to measure the interaction between a radioactively labeled ligand and a binding site, typically a receptor, transporter, or enzyme, under controlled conditions. Radioligands are ligands labeled with radioactive isotopes (e.g., tritium 3H, iodine-125, iodine-131, carbon-14, sulfur-35). The assay quantifies specific binding by separating bound from free ligand, often by filtration or precipitation, and detecting radioactivity with scintillation counting or gamma counters.

Two common assays: saturation binding to determine receptor density (Bmax) and affinity (K_d) by incubating membranes or cells with increasing concentrations of radioligand. Competition binding uses a fixed radioligand concentration with unlabeled competing ligand to estimate affinity (Ki) via the Cheng-Prusoff equation. Non-specific binding is defined in parallel with a saturating excess of an unlabeled ligand.

Key concepts: specific binding, nonspecific binding, receptor-ligand affinity, receptor density, and kinetics. Experimental considerations include maintaining receptor integrity, controlling temperature, ionic strength, and pH, and accounting for decay of radioactivity and radioligand stability.

Applications: pharmacology and neuroscience for characterizing receptors (adrenergic, dopaminergic, serotonergic), transporters, and enzymes; drug discovery and receptor pharmacology; autoradiography to map receptor distribution in tissue.

Limitations: use of radioactive materials requires safety protocols and regulatory approvals; in vitro conditions may not reflect in vivo physiology; radioligand decay and potential ligand-induced internalization can affect measurements.