A plaque-inhibition technique was used to study the response of mice to the dinitrophenyl (DNP) determinant at the cellular level. Sheep erythrocytes were sensitized for use in the plaque assay by exposing them to dinitrophenylated antisheep hemolysin of low hemolytic efficiency. DNP-lysine was added to the agar to inhibit specifically indirect plaques formed by mouse spleen cells producing anti-DNP antibodies. The average relative affinity of these antibodies was defined as the reciprocal of the molar concentration of DNP-lysine producing 50% plaque inhibition. The average relative affinity increased by a factor of 130 between 1 and 8 weeks after initial immunization. A highly significant correlation, apparently linear, was found between the average relative affinity and the average association constant of the serum antibody as measured by a modified Farr technique. The slopes of the plaque-inhibition curves in the vicinity of 50% plaque inhibition were assumed to be inversely proportional to the degree of heterogeneity of the antibodies with respect to affinity. There was a highly significant positive correlation between affinity and heterogeneity.

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