Use of protein-stained immunoblots for unequivocal identification of antibody specificities.

The unequivocal identification of immunochemically detected proteins on nitrocellulose blots requires precise alignment of the antibody-stained antigen spot with the protein-stained pattern. We report a method for the staining of proteins on nitrocellulose membranes immediately after electroblotting with Amido Black 10B under mild conditions which do not result in loss of antigenic activity. These membranes can then be probed with immunological detection reagents and the detected antigen correlated with the stained pattern. The procedure is simple, fast and particularly useful for the identification of antibody-labelled spots on blots of high-resolution two-dimensional gels.