For influenza H1N1 strains, including some of their escape variants, the association of amino acid differences located at their hemagglutinin HA1 domains with their antigenic relationship was examined. The antigenic relationship was recorded in terms of the ratios of hemagglutination inhibition (HI) titers, the concentration of antibody molecules recognized by the virus, and the equilibrium constant of epitope-paratope interaction determined with heterologous virus compared to that found with homologous virus. The HI titers of antisera were found to depend primarily on the concentration of antibody molecules recognized by the virus and much less on the equilibrium constants. The avidity of antibody in sera raised against historically later strains with earlier strains was higher than vice versa. In contrast to the results obtained with antisera, the same concentration of monoclonal antibody directed to the Sb site of A/Brazil virus was recognized by both heterologous and homologous viruses, and the differences in HI titers observed were due to avidity changes only. Some of the amino acid differences located at each of the antigenic sites were found to be associated with a reduction in the HI titers and in the concentration of antibody molecules recognized by heterologous virus, whereas other differences in addition decreased the avidity of epitope-paratope interaction. Further amino acid differences decreased the avidity only. The strains tested differed also in their amino acids located outside the antigenic sites. However, an influence of these differences on the reaction of virus with antibody could not be evidenced. For the strains tested, the antigenic hemagglutinin drift occurred by reduction of the concentration of antibody molecules recognized by the virus and by avidity changes, which, in turn, were caused by exchanges of some key residues located at the antigenic sites.