Two monoclonal antibodies directed to the blood group A determinant have been established. One hybridoma antibody (AH16), obtained on immunization with human gastric cancer cell MKN45, was directed to the blood group A determinant carried by both type 1 and type 2 chains. The other antibody (AH21) was prepared by immunization with a purified type 1-chain A glycolipid, followed by selection of the hybridoma by positive reaction with the type 1, but negative reaction with the type 2-chain A glycolipid. Thus, the antibody specifically reacts to the A determinant carried by the type 1 chain, but not to the A determinant carried by the type 2 chain; an A determinant carried by type 1 chain differs significantly from that carried by type 2 chain in its conformational structure. The antibody AH16 agglutinates both A1 and A2 human erythrocytes, but AH21 agglutinates neither A1 nor A2 erythrocytes, which supports the results of methylation analysis that blood group A and H determinants in human erythrocytes are exclusively made of the type 2 chain and that A1 and A2 phenotypes are not related to the difference between type 1 and type 2 chains.