Isotype Determination of Monoclonal Antibodies by Immunodiffusion

Immunodiffusion is an important qualitative method for the detection of antigens and antibodies. The technique may be used to determine the immunoglobulin heavy and light chain class and subclass of hybridoma antibodies. The monoclonal antibodies are allowed to diffuse toward anti-mouse isotype specific antisera in an agarose gel. In a positive reaction, a precipitin line will develop in the agarose between two wells. A monoclonal antibody should show only one heavy chain and one light chain. Knowledge of the isotype is also useful for purification purposes.