Affinity membrane identification of immunoglobulin subclass in hybridoma screening

Monoclonal hybridomas secrete immunoglobulins with a single antigen specificity and distinct class/subclass structure. Hybridoma management has commonly incorporated tests of antigen specificity into early screening procedures, but has not typically utilized assays of immunoglobulin structure. In this article, we describe a technique of class/subclass typing using polyvinylidene difluoride affinity membranes and a colorigenic enzymatic amplification system. The typing of monoclonal antibody structure was sufficiently sensitive to permit its routine use within several weeks of hybridoma fusion. The information obtained from early and routine class/subclass determinations included a semiquantitative assessment of monoclonal antibody concentration. In addition, the detection of a single immunoglobulin class/subclass in a microtiter well supernatant supported the possibility that the colony was monotypic. The application of class/subclass typing and Poisson statistics to hybridoma fusions provided a numerical estimate of the probability of colony monotypia.