The significance of multiple antibody components in serum of immunized rabbits.

The electrophoretic patterns of six sera from rabbits immunized by two or more courses of intravenous injections of killed pneumococci type III showed multiple peaks in the γ-globulin region. Such sera contained large amounts of antibody (up to 85 per cent of the total γ globulin) against the capsular polysaccharide. One serum contained a cryoglobulin, which contained almost as great a proportion of specific antibody as did the remaining γ globulin. The electrophoretic patterns and antibody contents were similar in the water-soluble and water-insoluble fractions of γ globulin. The sedimentation constant and diffusion coefficient of a water-soluble fraction of γ globulin, containing 85 per cent specific antibody, were measured. The values, at 0.4 per cent protein concentration, were S = 6.97×10 and D = 4.16×10 cm. sec., corresponding to molecular weight 159,000. The antibody-containing globulin from one serum was separated by zone electrophoresis into three fractions with different electrophoretic mobilities. These contained 53–71 per cent of antibody precipitable by type III pneumococcus capsular polysaccharide. Only doubtfully significant differences were found in respect of amino-acid composition, hexose and hexosamine contents, or antigenic characteristics. A method was devised for detecting small amounts of antibody against capsular polysaccharide by means of red cells sensitized with culture filtrates of capsulated pneumococci. The antibody was also fractionated by chromatography on anion-exchange cellulose, and numerous fractions with antibody activity were obtained. It was shown by labelling the γ globulin with I that similar fractionation occurred both in the presence and absence of other serum components. All the chromatographic fractions of γ globulin were found to contain approximately similar proportions of antibody. By electrophoresis in starch gel the fractions were found to differ from one another and to be heterogeneous. The implications are discussed of the finding that antibody against type III pneumococcus capsular polysaccharide can occur over the entire range of γ-globulin molecules.