Multiple immunoblot: a sensitive technique to stain proteins and detect multiple antigens on a single two-dimensional replica.
A multiple immunoblotting technique was developed to positively identify up to three different antigens on a single nitrocellulose replica of a two-dimensional isoelectric focusing/sodium dodecyl sulfate-polyacrylamide gel. Three highly sensitive immunoblot assays were selected, including: horseradish peroxidase/luminescence, alkaline phosphatase, and silver-enhanced immunogold. As a major advantage, the method permits a simultaneous detection of up to three different antigens without eluting the antibody-dye complex between staining of single polypeptides, thus providing a highly accurate identification of closely migrating components. The staining procedure is summarized in a flow chart. In addition to the multiple immunoblot staining, some suggestions are provided for a sensitive protein staining.
Steffen
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Department of Cell Biology and Neuroanatomy, University of Minnesota, Minneapolis 55455.
Linck
R W
RW
D000818:Animals; D000906:Antibodies; D000941:Antigens / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D015151:Immunoblotting / Q000379:methods*; D007150:Immunohistochemistry; D011506:Proteins; D012617:Sea Urchins; D013194:Staining and Labeling
D000906:Antibodies; D000941:Antigens; D011506:Proteins