Immunodetection of nitrocellulose-adhesive proteins at the nanogram level after trinitrophenyl modification.
A method for protein detection on nitrocellulose membranes based on modification with 2,4,6-trinitrobenzenesulfonic acid and reaction with anti-trinitrophenyl (TNP) serum as first antibody followed by peroxidase-conjugated second antibody is described. Protein quantities between 1 and 3 ng can be detected in the dot test. This method was used in a double immunodetection procedure after electrophoretic transfer of proteins localizing first a distinct antigen with its specific antiserum followed by visualization of the complete protein pattern on the same blot by the TNP/anti-TNP method as described above. As only water-soluble reagents are employed no shrinkage of the membrane occurs. Furthermore, the method can be used in a simultaneous immunodetection procedure visualizing the specific antigen together with TNP marker proteins using a mixture of the specific antiserum and the anti-TNP serum as first antibody.
D000818:Animals; D003101:Collodion; D004586:Electrophoresis; D007118:Immunoassay / Q000379:methods*; D009578:Nitrobenzenes; D011506:Proteins / Q000032:analysis*; D011817:Rabbits; D014302:Trinitrobenzenesulfonic Acid
D009578:Nitrobenzenes; D011506:Proteins; D014302:Trinitrobenzenesulfonic Acid; D003101:Collodion