Antibody Aggregate Removal by Hydroxyapatite Chromatography

Hydroxyapatite (HA) has proven in recent years to be one of the most versatile and powerful methods for removing aggregates from antibody preparations. It is effective with IgA, IgG and IgM, and it reduces aggregate levels from above 60% to less than 0.1%. Three basic elution strategies have evolved, one that removes aggregates from a modest proportion of clones, another from the majority, and one that appears to be universally effective. Each has distinct development and process ramifications. This review defines what HA is, how it interacts with various classes of biomolecules, how those interactions are controlled by different elution strategies, and how to determine which approach may be most effective for a particular antibody. Consideration is also given to HA's specific strengths and limitations from an industrial perspective.