A new and rapid method for immunoglobulin class and subclass determination of mouse monoclonal antibodies using a solid-phase immunoradiometric assay

Abstract

A solid-phase immunoradiometric assay is described for the detection of mouse immunoglobulin classes and subclasses in unpurified and unconcentrated supernatants of hybridomas. IgG fractions from rabbit antisera specific for mouse immunoglobulin classes and subclasses are used for coating the wells of flexible microtiter plates. Monoclonal antibody present in hybridoma supernatants is bound only to wells that contain the appropriate anti-subclass antibody. The binding of hybridoma antibodies to corresponding IgG subclasses or IgM is then detected by a labeled rabbit anti-mouse antibody binding to all mouse immunoglobulins (heavy and light chains). Thus, only 1 labeled antibody is needed for all assays. The advantages of the method described are the following: results are obtained within a few hours and antibody containing hybridoma supernatants may be used without a concentration step since minute amounts of antibody are detected by the immunoradiometric assay. Cultures producing several subclasses may be early recognized as oligo/polyclonal.