Multiple Peak Formation of Avidin on an Iminobiotin Affinity Support

When pure avidin is chromatographed on a homemade iminobiotin column, two distinct peaks are observed using a gradient solvent system starting with a pH 9 buffer. Proteins in both peaks are shown to be identical by nondenatured polyacrylamide gel electrophoretic experiment. The weak interaction of the protein with the stationary phase at pH 9 could cause this unusual behavior.