A method for the determination of antibody affinity using a direct ELISA

Antibody affinity is of major significance in immunoassays. Since affinity may be influenced by the immunoassay methodology it is important to determine this parameter under the conditions of the assay used. Here a method is described for the determination of binding constants (K) in a direct ELISA with the use of the computer program LIGAND. Five of the antibodies studied bound to their antigen with two classes of antigen binding site, while all the other antibodies studied reacted with only a single class of antigen binding site. The accuracy of the method and the implications for atigen-antibody reactions are discussed.