Isothermal titration calorimetry (ITC) is used in thermodynamic analyses of macromolecular interactions. This technique is often referred to as a true in-solution method, as it does not require chemical tagging or immobilization of the interacting components.  Common applications include analysis of protein interactions with another protein, peptide, metal, or nucleic acid.  In addition, ITC can be used to examine activity of enzymes.

The measured heat, either absorbed or released upon binding, provides the basis for determination of:

• binding constants (Ka)
• enthalpy (ΔH)
• entropy (ΔS)
• stoichiometry of the reaction
• enzyme kinetics parameters

Instrument Specs

The MicroCal VP-ITC is a highly sensitive calorimeter that allows for analysis of dilute solutions of macromolecules (typically in the micromolar range for proteins). The instrument sample cell volume is approximately 1.5 mL, with the injection syringe containing about 290 µL of titrant. The isothermal operation is carried out within the temperature range of 2^oC to 80^oC.

More information

See vendor's site: MicroCal

Suggested Reading

• Reference List at MicroCal
• Leavitt S. & Freire E. (2001) Direct Measurement of Protein Binding Energetics by Isothermal Titration Calorimetry, Curr. Opin. Struct. Biol., 11, 560-566.
• Ladbury J.E. & Chowdhry B.Z. Eds (1998) Biocalorimetry: Applications of Calorimetry in the Biological Sciences, John Wiley and Sons.
• Harding S.E. & Chowdhry B.Z., Eds (2001) Protein-Ligand Interactions: Hydrodynamics and Calorimetry, A Practical Approach, Oxford University Press.
• Cooper A., Johnson C.M., Lakey J.H. & Nollmann M. (2001) Heat Does Not Come in Different Colours: Entropy-enthalpy Compensation, Free Energy Windows, Quantum Confinement, Pressure Perturbation Calorimetry, Solvation and the Multiple Causes of Heat Capacity Effects in Biomolecular Interactions, Biophys. Chem., 93, 215-230.
• Jelesarov I. & Bosshard H.R. (1999) Isothermal Titration Calorimetry and Differential Scanning Calorimetry as Complementary Tools to Investigate the Energetics of Biomolecular Recognition, J. Mol. Recognit., 12, 3-18.
• Read C.M. & Jelesarov I. (2001) Calorimetry of Protein-DNA Complexes and Their Components, in Methods in Mol. Biol., Moss, T., Ed., 148, 511-533.
• Ladbury J.E. (2004) Application of isothermal titration calorimetry in the biological sciences: things are heating up! Biotechniques, 37, 885-7.
  Perozzo R., Folkers G., & Scapozza L. (2004) Thermodynamics of protein-ligand interactions: history, presence, and future aspects.
  J. Recept. Signal. Transduct. Res., 24, 1-52.
• Velazquez-Campoy A., Leavitt S.A., & Freire E. (2004)Characterization of protein-protein interactions by isothermal titration calorimetry. Methods Mol. Biol., 261, 35-54.
• Lewis E.A. & Murphy K.P. (2005) Isothermal titration calorimetry.
  Methods Mol. Biol.,3 05, 1-16.
• Morin, P.E. & Freire, E. (1991) Direct calorimetric analysis of the enzymatic activity of yeast cytochrome c oxidase, Biochemistry 30, 8494-8500.
• Todd M.J. & Gomez J. (2001) Enzyme kinetics determined using calorimetry: a general assay for enzyme activity? Anal. Biochem., 296, 179-87.

ITC protocols and procedures

Any manuals posted here are EXCLUSIVELY for the use related to the instruments housed in the BPL. All copyrights apply.