Application of the nitrocellulose transfer technique and alkaline phosphatase conjugated anti-immunoglobulin for determination of the specificity of monoclonal antibodies to protein mixtures.

The target antigens of a series of monoclonal antibodies raised against a complex mixture of proteins have been identified. The protein mixture was fractionated by polyacrylamide gel electrophoresis in SDS, and the fractionated proteins electrophoretically transferred to nitrocellulose sheets. Protein bands reacting with antibody were identified by incubating the nitrocellulose strips with the 'hybridoma' culture supernatants, followed by detection of bound antibody using alkaline phosphatase conjugated to rabbit anti-mouse F(ab')2 antibody, and histochemical substrates. With this technique, the antigens reacting with 4 of a series of 5 monoclonal antibodies have been identified.