Modified electrode buffers for stain-free protein detection in electrophoresis

Abstract

Proteins that are electrophoretically separated in a gel are derivatized to produce fluorescent emissions by incorporating halo-substituted organic compounds into one or both of the electrode buffer solutions at the two ends of the gel. The halo-substituted compounds used are ones that bear an electric charge at the pH of the buffer solutions and gel, and the polarity of the charge on the compounds is such that the compounds migrate from the electrode buffer into the gel under the electrophoretic influence concurrently with the migration of the proteins into the gel. Once the proteins are separated and distributed within the gel and the gel is fully penetrated with the halo-substituted compounds, the gel is irradiated with ultraviolet light to induce a reaction between the halo-substituted compounds and the proteins through the tryptophan residues on the proteins, producing fluorescent reaction products.