ITC (Isothermal Titration Calorimetry) Analysis

ITC (Isothermal Titration Calorimetry) is a direct, label-free biophysical technique used for real-time monitoring and quantification of intermolecular interactions. Its core principle is as follows: at a constant temperature, one reactant is added dropwise to a sample cell containing another reactant using a syringe, and the trace amounts of heat released or absorbed during each titration due to molecular binding or dissociation are measured in real time. The instrument maintains the temperature of the sample cell and the reference cell at the same level through a power compensation system; the compensation power is a direct measure of the heat of reaction. Integrating the thermal peak of each injection yields the binding isotherm, and then, through nonlinear fitting, complete thermodynamic parameters such as binding affinity (KD), stoichiometry (n), enthalpy change (ΔH), entropy change (ΔS), and Gibbs free energy (ΔG) are obtained.