A convenient enzyme-linked immunosorbent assay for testing whether monoclonal antibodies recognize the same antigenic site. Application to hybridomas specific for the beta 2-subunit of Escherichia coli tryptophan synthase

Abstract

Seven hybridoma clones, producing antibodies directed against the beta 2-subunit of Escherichia coli tryptophan synthase, have been obtained from mouse cells. To test whether the corresponding monoclonal antibodies recognize different epitopes on beta 2, an ELISA double antibody binding system has been developed and is reported here. The antigen is first coated onto a microtitration plate. Two monoclonal antibodies are then added either separately or simultaneously, and the amount of bound antibody is quantitatively measured by use of immunoglobulin (rabbit anti-mouse IgG) linked to beta-galactosidase. Additivity of the bound enzymatic activity is observed when the monoclonal antibodies bind to distinct epitopes. Using this test, it is shown that, of the 7 anti-beta 2 monoclonal antibodies obtained, 5 recognize the same antigenic site and the 2 others recognize a second site.