Biophysical Analysis

Isothermal Titration

Isothermal titration calorimetry (ITC) is a sensitive method for characterizing the binding between two molecules. The method results in determination the binding constant and stoichiometry of binding, as well as other thermodynamic parameters such as the molar heat, free energy, and entropy of binding. The method is based on the principle that binding of two molecules results in heat being given off (an exothermic event), or heat being required (endothermic). In an ITC experiment, one component (e.g. an antibody) is placed in a stirred cell thermostatted to a constant temperature. Incremental injections of the ligand (e.g. an antigen) are made into the cell. After each injection, the amount of heat given off (or required) to maintain the constant temperature is measured. In this way, a titration curve is generated from which the stoichiometry and equilibrium binding constant can be calculated.



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