A general immunochemical method for detecting proteins on blots.
Following horizontal electroelution, or blotting, of proteins from polyacrylamide gels to immobilizing matrices, such as nitrocellulose or Zeta-bind paper, the transferred proteins can be derivatized in situ with pyridoxal 5'-phosphate and sodium borohydride. After a quenching step to eliminate nonspecific binding of antibody to the protein-binding matrix, the blot is incubated with a solution containing a mouse monoclonal antibody specific for the 5'-phosphopyridoxyl group. The transferred proteins can then be located on the blot with second antibody staining procedures employing either a peroxidase-linked goat anti-mouse F(ab')2 antibody or a peroxidase-linked avidin/biotin system. The solid-phase enzyme-linked immunosorbent assay method described in this report is a mild, general, and sensitive immunochemical method for the detection of proteins on protein-binding matrices.
D000818:Animals; D000911:Antibodies, Monoclonal; D002843:Chromatin / Q000302:isolation & purification; D003101:Collodion; D003600:Cytosol / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay; D006657:Histones / Q000302:isolation & purification; D007124:Immunoenzyme Techniques; D008099:Liver / Q000032:analysis; D009757:Nylons; D010209:Paper; D011506:Proteins / Q000032:analysis* / Q000302:isolation & purification; D011732:Pyridoxal Phosphate; D051381:Rats
D000911:Antibodies, Monoclonal; D002843:Chromatin; D006657:Histones; D009757:Nylons; D011506:Proteins; D011732:Pyridoxal Phosphate; D003101:Collodion