(141b) A New Multimodal Membrane for Monoclonal Antibody Purification

Authors: 
Wang, J., Clemson University
Husson, S. M., Clemson University

This presentation will describe our latest research efforts to develop a new class of multimodal membranes (MMM) with high binding capacities, salt-tolerance, and high-productivity for the chromatographic purification of therapeutic proteins. The membranes incorporate ligands that can bind proteins through a combination of Coulombic interactions, hydrophobic interactions, and/or hydrogen bonding, thus widening the operation ranges for separation.

Our design approach is to graft functional polymer tentacles from the pore surfaces of macroporous membranes using a controlled surface-initiated polymerization technique. One advantage of this membrane preparation technique is that it greatly enhances the membrane binding capacity without severely decreasing its permeability. Here, we present work on the preparation of multimodal cation-exchange membranes for bind-and-elute purification of immunoglobulin G (IgG). Compared to commercial multimodal adsorbents, our membranes have 4 times higher ligand density, which contributes to the high static binding capacities (180 mg IgG/mL, 310 mg lysozyme/mL).

Protein binding experiments with bovine IgG showed binding capacities of the multimodal membranes were dependent on pH but had very good salt tolerance. The multimodal membranes maintain significant binding capacities in excess of 90 mg IgG/mL at ionic strength values that are typical for elution buffers used in multi-stage bioseparation processes.  For sodium citrate, a conventional salt used in elution buffers of Protein A columns, increasing ionic strength had only a minor effect on the IgG binding capacity.

To further understand protein adsorption on the multimodal membrane, a thermodynamic adsorption isotherm model was employed that provides a unique set of physically meaningful parameters. This model can be used to explain the effect of salt type (kosmotropes, neutral, and chaotropes) on protein adsorption.

Finally, protein dynamic binding capacities were determined from breakthrough curve analysis. Dynamic binding capacities of >50 mg IgG/mL were found near the isoelectric point of the protein. A range of flow rates was used to study the effect of volumetric throughput on dynamic binding capacities. Protein elution using pH adjustment, chaotropic salts, and organic displacers will be described.

Taken together, the results to be presented indicate that the newly developed multimodal membranes have great potential to compete with more traditional cation-exchange materials following the Protein A purification step in the downstream processing of antibody products.

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