DETERMINATION OF THE CONCENTRATION OF INFLUENZA-VIRUS ANTIHAEMAGGLUTININ ANTIBODY MOLECULES OF THE IGG CLASS AND OF THE EQUILIBRIUM-CONSTANT BY USE OF ENZYME-IMMUNOASSAY TITERS DETERMINED FOR GRADED EPITOPE CONCENTRATIONS

A new technique for determining the concentration of influenza virus a ntihaemagglutinin antibody molecules of the IgG class (A) and of the e quilibrium constant K of paratope-epitope interaction is described. Th e method is based on determining enzyme immunoassay (EIA) titres of an tibody with graded epitope concentrations: EIA titres were defined in terms of the antibody dilution yielding a fixed amount of antibody ads orbed per ml (= 6.21 X 10(10)). Adsorption of antibody depends on the concentration of paratopes and epitopes allowed to react and on the eq uilibrium constant. For the use of a constant concentration of epitope s, the paratope concentration needed to yield the desired degree of an tibody adsorption decreases with increasing avidity. Therefore, the EI A titres increase both with increasing avidity and increasing antibody concentration. When graded epitope concentrations are used for determ ining the EIA titres of a given serum, the titres are influenced in a similar manner by the antibody concentration of the serum and the incr ease of titres with increasing epitope concentration reflects avidity. The equilibrium constant found is subsequently used to determine the concentration of free antibody at the dilution meeting the definition of EIA titre and the product of EIA titre and the sum of free and boun d antibody at this dilution gives the number of antibody molecules pre sent in the test serum. A panel of 118 antisera was tested comparative ly for A and K using the new method and by means of the guanidine titr e ratio test and equilibrium filtration. The values obtained agreed we ll with each other. This novel technique offers the advantage that it can be easily adapted for use with other viruses.