Chemical Resolution of an Epitope Recognized by Blood Group Antibodies Capable of Binding Both A and B Red Blood Cells

The high specificity of human antibodies to blood group A and B antigens is impressive, especially when considering the structural difference between these antigens (tetrasaccharides) is a NHAc versus a hydroxyl group on the terminal monosaccharide residue. It is well established that in addition to anti‐A and anti‐B there is a third antibody, anti‐A,B capable of recognizing both A and B antigens. To analyze this AB specificity, we synthesized a tetrasaccharide, where the NHAc of the A antigen was replaced with an NH2. This NH2 group was then used to attach the glycan to an affinity resin, creating an AB epitope (ABep) adsorbent where the critical site for recognition by A and B antibodies was not accessible, while the rest of the (conformationally compact) tetrasaccharide remained accessible. Anti‐ABep antibodies were then isolated from blood group O donors and found to have expected A,B specificity against immobilized and red cell bound synthetic antigens, including ABep, and were able to agglutinate both A and B red cells. The amount of these anti‐ABep (anti‐A,B) antibodies found in the blood of group O donors was comparable to levels of anti‐A and anti‐B found in group B and A individuals. Using STD‐NMR the location for the AB epitope on the tetrasaccharide was found. Antibodies were isolated from the blood of group O individuals that bound equally to antigens A and B, including agglutination of erythrocytes both A and B. STD NMR data correspond to the topography of the predicted epitope: STD‐active H atoms, shown in red, are concentrated at the same epitope subsite.