8] Antiisotypic antibodies

Abstract

This chapter provides an overview of antiisotypic antibodies. Antibodies capable of specifically binding to all molecules of an immunoglobulin class have proven to be invaluable reagents for a variety of purposes. This chapter reviews principles of preparation, characterization and application of antibodies. Sera (or ascites) containing substantial concentrations of a homogeneous Ig protein of the desired isotype (example, myeloma and hybridoma) are particularly advantageous starting materials for isolation of human, murine, and rat Ig proteins. Purified Ig proteins from several species are also available commercially. Regardless of the source, the purity of the proteins to be utilized for immunization should be ascertained by disc gel electrophoresis and immunoelectrophoresis against antiserum directed against serum proteins of the appropriate species prior to use. Enrichment of antiisotypic antibodies from heterologous antisera (or ascites/culture supernatant in the case of monoclonal antibodies) is frequently desirable prior to utilization. For heterologous antisera, either isolation of the IgG fraction of the serum or affinity purification of the antiisotypic antibody is usually required. Monoclonal antiisotype antibodies can generally be isolated from ascites (or culture supernatant) using affinity purification techniques. The chapter further discusses affinity purification of antiisotypic antibodies.