Insoluble blood group substances prepared by copolymerization of soluble blood group substances with N-carboxy-L-leucine anhydride were used to absorb blood group antibodies from two human, high-titered anti-A sera. After the absorbants were washed free of nonspecific serum proteins, blood group antibodies were eluted either with pH 3.62 acetate buffer, or at neutral pH with monosaccharide haptens of the A or B antigenic determinants (N-acetyl-D-galactosamine or D-galactose respectively). The purified anti-A antibodies were characterized, immunoelectrophoretically, as γM-, γA-, and γG-immunoglobulins. These were further separated into γM- and γG-fractions by gel filtration or density gradient centrifugation. Both γM- and one of the two γG-antibody fractions contained K and L light chain determinants; the remaining γG-fraction was comprised, almost totally, of type K molecules.

Precipitability of the purified anti-A immunoglobulins by blood group A substance varied from 43 to 89%. The agglutinating activity per unit N of the isolated γG-anti-A was found to equal, in one case, and to exceed, in the second, that of the γM-antibodies from the same individuals.

The marked differences between γM- and γG-antibody fractions in quantitative hapten inhibition studies were interpreted to mean that the antibody-combining site of the isolated eluted γG-anti-A was significantly larger than that of the eluted γM-anti-A. Whether these data connote differences in combining site size between entire immunoglobulin classes in an individual serum or simply reflect the properties of highly selected antibody populations cannot be stated at present.

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