Method for determining the concentration of influenza virus antihemagglutinin antibody molecules, the number of epitopes per virus particle and the equilibrium constant of virus antibody interaction

Abstract A novel technique for the determination of the concentration of influenza virus antihemagglutinin antibody molecules ( A ), of the equilibrium constant ( K ) and of the number of epitopes recognized per virus particle ( s ) is described. The test utilizes measurements of virus-antibody interaction both in a photometric hemagglutination inhibition test and in the method of equilibrium filtration and is based on the following principle: the number of antibody molecules adsorbed per virus particle at the antibody dilution ( 1 d 50 ) yielding 50% hemagglutination inhibition in a photometric HI test was found to be constant, providing an estimate of the concentration of bound antibody at the dilution d 50 . The corresponding concentration of free antibody is calculated by use of parameters determined by equilibrium filtration and the antibody concentration is obtained as sum of bound and free antibody. The antibody concentration found is used for calculating the equilibrium constant K and the number of epitopes recognized per virus particle.