Interaction of Protein Antigens and Antibodies

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Summary Experiments have been conducted on the solubilization and dissociation of specific diphtheria toxoid-antitoxin precipitates by acid and alkaline solutions and by alkaline solutions containing urea and thiocyanate. Additions of urea and thiocyanate were observed to have synergistic effects in solubilizing specific precipitates. Diphtheria toxoid-antitoxin precipitates could readily be dissolved under conditions which were sufficiently mild to prevent loss of serological activity of both components exposed separately, for example, in 0.1 M glycine buffer, pH 11.0, 2 C. Electrophoretic analyses of such dissolved precipitates showed that they migrated essentially as single components. More drastic conditions resulted in slight separation of the precipitates as measured by electrophoretic analyses. However, diphtheria toxoid and antitoxin exposed separately to these more drastic conditions showed some denaturation as demonstrated by decreased precipitate formation or complexing. These experiments support the suggestion that the forces involved in the interaction of protein antigens and antibodies are similar in nature and magnitude to those responsible for maintaining the native structure of the proteins. Diffusion experiments with a diphtheria toxoid-antitoxin precipitate dissolved at 2 C in 0.1 M glycine buffer, pH 10.8, containing 1 M urea and 0.1 M KCNS showed that the dissolved precipitate exists under these conditions as large, undissociated aggregates. This result is most readily explained by the theory that antibodies are bivalent. The results reported together with other data are interpreted as in harmony with the view that successive specific interactions between antigens and antibodies are accompanied by a decrease in affinity so that interactions between aggregates to give precipitates are weaker than the primary interactions between antigens and antibodies.