THERMODYNAMIC PARAMETERS IN IMMUNOASSAY

Although affinity and kinetic measurements on their own provide useful information regarding the suitability of antibodies for various immun oassays, a thermodynamic analysis provices additional information that throws light on the molecular forces at work in the antigen-antibody interaction. It may then be possible to adjust assay conditions in ord er to favour either the association or dissociation of antigen-antibod y complexes. A tenfold increase in binding affinity (K) corresponds to a free energy change of only 1.4 kcal/mol (5.8 kJ/mol) at 25 degrees C. This means that K values of 10(5) M-l and 10(10) M-1 correspond to a free energy change (Delta G) of 7.0 and 14.0 kcal/mol respectively. The entire range of affinity constants normally encountered in antigen -antibody interactions, therefore differs by no more than about 7 kcal /mol of free energy change, which is equivalent to only a few hydrogen bonds. In comparison, a single electrostatic interaction corresponds to about 4 kcal/mol of free energy change. A full description of the b inding interaction requires an understanding of the change in hydratio n states of the reactants when the complex forms, and an assessment of the entropic andenthalpic effects of these changes. Contrary to earli er assumptions, it is now clear that antigen-antibody interactions are often accompanied by a large favourable enthalpy which more than comp ensates the unfavourable entropy.