Both a new method for measuring protein ion exchange kinetics and new data for the adsorption of three different proteins in anionic agarose gels are described. The gels were prepared as spherical particles to determine chemical and structural properties and the macroscopic protein adsorption behavior, and as thin slabs supported in a microfluidics chip to observe transient concentration profiles with a light microscope. The combination of these approaches provides deeper insight in the nature of protein transport mechanisms than previously possible. The adsorption of cytochrome c, myoglobin, and hemoglobin was studied macroscopically and microscopically. Although all proteins exhibited similarly favorable adsorption isotherms, adsorption kinetics, and concentration profiles were quantitatively and qualitatively different. Cytochrome c was adsorbed very quickly and resulted in diffuse concentration profiles, while myoglobin adsorbed much more slowly and resulted in sharp fronts. Hemoglobin exhibited lower rates with diffuse profiles at higher pH and sharp fronts at lower pH. In general, the shape of the profiles appeared to be correlated with the binding strength. Reversibly bound proteins yielded diffuse profiles, while irreversibly bound proteins exhibited sharp advancing fronts, indicating that protein-surface interactions play a major role in the nature of the transport mechanism. (c) 2007 American Institute of Chemical Engineers