Equine Anti-Hapten Antibody

Abstract

Summary Antibody specific for the Lac haptenic group has been purified by a method of coprecipitation from the serum of a horse during a course of immunization with Lac-conjugated proteins extending over a year. The antibody was studied and found to contain at least six antigenically distinct immunoglobulins with the major components identifiable as γG and γA antibodies. DEAE-cellulose chromatography was employed to separate three major fractions containing mainly γGa, b, γGc and γA respectively. These isolated immunoglobulins were shown to have properties consistent with those described for these immunoglobulins in other species including electrophoretic mobility, sedimentation properties, and carbohydrate content. Their antigenic individuality was demonstrated to be a characteristic of their heavy chains. The antibody activities of these fractions were compared and marked differences were shown in the reactivities of the anti-Lac antibody populations of differing immunoglobulin content. The γGa, b antibody preparation precipitated and fixed complement with antigen; the γA antibody did neither and γGc was shown incapable of fixing complement. In sera collected early in the course of immunization the γG antibodies bound Lac dye with an association constant of < 5 × 105 M-1 while the γA had an affinity of > 107 M-1. As immunization progressed, however, the affinity of the γA remained fairly constant while that of the γG increased to > 107 M-1. Studies of the serum and total anti-Lac antibody preparation indicated that the properties of the serum reflected strongly the properties of the major classes of antibody it contained and that changes in the relative amounts or reactivities of these classes caused changes in the properties exhibited by the antisera.