The University of Massachusetts Amherst

Malvern Auto-iTC200

The MicroCal Auto-iTC200 is a fully automated (cleaning steps, titrations, filling cells, equilibrations, etc.), low volume, highly sensitive isothermal titration calorimeter. It delivers direct, label-free, in solution measurement of all binding and thermodynamic parameters in a single experiment. Applications include characterizing molecular interactions of small molecules, proteins, antibodies, nucleic acids, lipids and other biomolecules. It can also be used to measure enzyme kinetics.

Principles of Isothermal Titration Calorimetry (ITC)

Isothermal titration calorimeters measure the heat change that occurs when two molecules interact. Heat is released or absorbed as a result of the redistribution and formation of non-covalent bonds when the interacting molecules go from the free to the bound state. ITC monitors these heat changes by measuring the differential power, applied to the cell heaters, required to maintain zero temperature difference between the reference and sample cells as the binding partners are mixed.

Schematic representation of the essential components of an ITC system

The reference cell is usually filled with water. The sample cell contains one binding partner and a stirring syringe which holds the ligand. The ligand is injected into the sample cell, in small aliquots, until the ligand concentration is 2-3 times greater than the sample. Each injection of ligand results in a heat pulse that is integrated with respect to time and normalized for concentration to generate a titration curve of kcal/mol vs molar ratio (ligand/sample). The resulting isotherm is fitted to a binding model to generate the affinity (KD), stoichiometry (n) and enthalpy of interaction (ΔH).