Adsorption of deamidated antibody variants on macroporous and dextran-grafted cation exchangers: I. Adsorption...

Single and multicomponent adsorption isotherms were obtained for deamidated variants of a monoclonal antibody on two cation exchangers with different pore structures, one with a macroporous architecture--UNOsphere S, and the other with charged dextran grafts--Capto S. No selectivity between the different deamidated forms was seen at pH 5 for either stationary phase. However, although the binding strength was lower, both media exhibited substantial selectivity at pH 7.5. The effective binding charge, determined from linear gradient elution experiments and from the steric mass action model, was different for the two different media, but remained nearly the same for the different variants suggesting that the selectivity is determined by the strength of binding, rather than by the binding charge. This result agrees with the higher binding constants determined for the less deamidated forms. At low ionic strength, the binding capacity of the dextran grafted media was much higher than that of the macroporous matrix at either pH. However, similar capacities were obtained for the two stationary phases at ∼140 mM Na+ for pH 5 and at 50 mM Na+ for pH 7.5. For both materials, predictions of multicomponent adsorption based on the steric mass action model were in good agreement with experimental results indicating that the different variants bind in competition with each other. In general, this work demonstrates the utility of modeling adsorption equilibrium allowing an accurate description of competitive binding, which is a necessary step for a complete description of a chromatographic separation process.