pH and conductivity transients during elution of IgG from protein A columns.

Abstract

The evolution of pH and conductivity waves during elution of IgG from protein A columns is studied for the resin MabSelect PrismA as well as other commercial resins using glycine and acetate buffers as eluents. The effects of buffer composition, IgG load, and residence time are explored. For glycine buffers, conductivity and pH waves travel through the column at different speeds, with the conductivity wave emerging in the column void volume and the pH waves emerging in 1 to 6 column volumes (CV) dependent on the buffer composition. The pH effluent temporarily overshoots the feed value, followed by a sharp drop as the pH approaches the eluent pH. For these conditions, elution of IgG is delayed and, at high loads, most of the IgG loaded elutes from the column at high pH values. Complex pH profiles, involving overshoots and delays between conductivity and pH waves are also observed when eluting with dilute sodium acetate (50 mM) or with acetic acid, but to a lesser extent. No overshoots or delays are observed for more concentrated sodium acetate (100 mM). A mechanistic model is developed to predict elution with glycine buffers. Model predictions agree with the experimental trends. In particular, the model shows that when eluting a partially loaded column, IgG desorbed near the column entrance is re-adsorbed downstream of the pH front eventually emerging at high concentration and high pH. The model can be used to design optimized buffers and predict the pH of the IgG elution pool.