The removal of aggregated species, particularly soluble dimers, from monoclonal antibodies (mAbs) remains a persistent challenge in downstream processing. In this work, we have examined the separation of a mAb monomer and of its dimer on the cation exchange resin Nuvia HR-S (BioRad Laboratories) using frontal analysis. In such a process, a mixture of monomer and dimer is continuously fed to the column under conditions where the mixture is strongly adsorbed. Separation is achieved through competitive binding mechanisms, resulting in a series of fronts whose monomer and dimer composition are determined by the multi-component equilibrium and kinetics of the system. Experimentally, the selectivity for dimer was found to vary substantially with ionic strength, being lowest when conditions favor the strongest binding, and increasing to a maximum at intermediate ionic strengths where surface coverage is lower. A large kinetic resistance to exchange of the monomer and dimer at low ionic strengths is found to be responsible for this behavior. Thus, the intrinsic kinetics of adsorption and desorption for the monomer and dimer on ion-exchange surfaces are investigated at various ionic strengths using biolayer interferometry. Finally, a computational model is developed to describe the competitive binding frontal analysis process, accounting for process parameters such as feed composition and salt concentration, which can be used to optimize column operating conditions for binding capacity and selectivity.

Competitive binding kinetics of antibody monomer-dimer mixtures on CEX resins during separation by frontal analysis

253rd National Meeting (2017)

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