1. Fluorescence quenching has been calibrated, by comparison with equilibrium dialysis, for measurement of antibody-hapten association constants with guinea pig antibody to the 2,4-dinitrophenyl (DNP) determinant. The maximum quenching with all sites occupied which gave best agreement with equilibrium dialysis was found to be 100%.
2. The carrier to which the DNP-hapten is coupled influences significantly the amount, class (γ1 or γ2), and affinity of the antibody of DNP specificity made by individual guinea pigs.
3. The affinities of γ1- and γ2-antibodies from individual guinea pigs are generally very similar.
4. Precipitin curves with γ1-antibodies show more marked inhibition of precipitation in antigen excess than do precipitin curves with the less charged γ2-antibodies, indicating the importance of nonspecific forces in the precipitation reactions.