A Tool Box of Advanced Sorbents for High Productivity Chromatography

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Biologic APIs are becoming more complex, and upstream production processes are generating higher-titer process streams, increasing the demand for more efficient downstream purification solutions. Because proteins are complex natural molecules, each biologic drug substance interacts with chromatography sorbents in a unique manner. Rapid and thorough screening of chromatography sorbents as early as possible in the process development phase is therefore necessary to identify an economical and operationally efficient combination for a given biologic API.
In the following application note, Pall Life Sciences provides an overview of a toolbox of highly selective and robust (for both clean-in-place [CIP] and single-use operations) scalable chromatography sorbents and membrane separation devices to support high productivity, large-scale batch or continuous chromatographic separations of biologic APIs. It is important to note that all portfolio options are also available in formats to support high-throughput chromatography sorbent screening and optimization.

HyperCel™ ion exchange sorbents
The newest ion exchange sorbents in the Pall Life Sciences portfolio, HyperCel STAR AX and HyperCel STAR CEX, have been designed to provide excellent separation to streamline downstream processing under the higher salt conditions commonly observed today; they are ideally suited for capture, intermediate, and polishing steps in the protein purification process. Both sorbents offer flexibility for the use of higher conductivity feed streams and limit the need for intermediate diafiltration and/or dilution.

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HyperCel STAR AX, a scalable anion exchange sorbent designed for high-productivity protein capture at moderate or higher salt conductivities (2-15 mS/cm), can bind proteins under close to physiological conditions, allowing the direct use of most bioprocess fluids without dilution. This ability to directly capture proteins from undiluted feed streams means that HyperCel STAR AX sorbent can be used for early contaminant removal prior to final purification.
HyperCel STAR CEX is a scalable cation exchange sorbent designed for high productivity protein capture and impurity removal at moderate salt conductivities (2-12 mS/cm) – conductivities slightly higher than those applied with classical cation exchangers.
With rigid cellulose matrices, both HyperCel STAR AX and STAR CEX are suitable for large-scale processing of fermentation and cell culture feedstocks, offering flow properties and backpressures compatible with the needs of commercial-scale protein production. They also exhibit high dynamic binding capacities at short residence times, allowing for moderate column volumes and lower footprints, which translate to reduced buffer volumes and consequent cost savings. The chemical stability of these ion exchange sorbents allows for CIP with 0.5 to 1 M sodium hydroxide (NaOH) and long-term storage in 10 to 100 mM NaOH.

HyperCel mixed-mode sorbents
Mixed-mode or multi-mode chromatography exploits different ligand-protein interaction mechanisms and provides new selectivities for protein separations that are based on a combination of ion exchange and hydrophobic interactions. The mixed-mode sorbent family includes anion exchange sorbents MEP HyperCel, HEA HyperCel, and PPA HyperCel (all of which contain a positively charged moiety) and cation exchange sorbent CMM HyperCel (which contains a negatively charged moiety). All are based on robust synthetic ligands immobilized on an industry-scalable cellulose matrix.
MEP HyperCel sorbent is immunoglobulin-selective and designed for capture and purification of antibodies and various recombinant proteins from process development to manufacturing scale. As a hydrophobic charge induction chromatography sorbent, MEP HyperCel has a unique separation mechanism that is a no-salt/low-salt alternative to hydrophobic interaction chromatography (HIC) sorbents with simplified process operations and enhanced process economics. It also allows for the separation of proteins with similar isoelectric points.
HEA and PPA HyperCel mixed-mode sorbents also function on the basis of a combination of electrostatic and hydrophobic interactions between proteins and the sorbent ligands. Each product offers unique selectivities that are different from those of ion exchange or conventional HIC sorbents. They are ideal for separating challenging mixtures of proteins and, due to high mechanical stability, can be used at high linear velocities at both process development and commercial scales.
The recently introduced CMM HyperCel sorbent is a scalable, negatively charged mixed-mode sorbent for high performance capture and impurity removal at moderate salt conductivities, particularly from challenging process fluids. It directly complements the positively charged MEP, HEA and PPA HyperCel mixed-mode sorbents. CMM HyperCel separates proteins with similar isoelectric points and/or hydrophobicity at low or high conductivity, retains its high dynamic binding capacity over repeated purification cycles, provides high recovery yields with low elution volumes and is easy to regenerate.

KANEKA™ KanCapA Protein A sorbent
KANEKA KanCapA is a state-of-the-art Protein A chromatography sorbent from Kaneka Corporation Japan. Comprised of a proprietary recombinant Protein A ligand covalently bound to a rigid cellulose matrix, KANEKA KanCapA chromatography sorbent is readily scalable, easy to pack, has a high binding capacity to match current expression levels and high selectivity under mild conditions; furthermore, it maintains this performance over long service periods. It excels in primary capture of monoclonal antibodies from clarified cell cultures.

Mustang® membrane chromatography devices
With an open pore structure and a portfolio of Q, S and E surface chemistries, Mustang membrane chromatography eliminates the diffusion limitations of sorbents, allowing biomolecules to access all binding sites by direct fluid convection. This means that high dynamic binding capacities and sharp breakthrough curves are achieved over a very wide range of flow rates and also a large range of molecular sizes. Mustang ion exchange membrane chromatography devices offer a flexible solution in bioprocessing, with scalable devices that can be used either as single- or multiple-use products. High volumetric flow rates lead to increased throughput, and the simple, easy-to-use compact design enables reduced buffer consumption and overall process economic gains. Mustang membrane products are ideal for efficient contaminant removal and capture of large target molecules.

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Mustang Q, S, and E capsules are disposable, single-use devices available in a range of volumes. Mustang Q and S membranes have, respectively, anion and cation exchange properties that afford high dynamic capacities for macromolecules including DNA, plasmids, Factor VIII, viral vectors and viruses. Mustang E membrane embraces a positively charged polymer that is suited for the removal of endotoxin from process fluids such as buffers and water. The products in the scalable Mustang Q XT capsule range can also be used as disposable devices but have been designed with a robust polypropylene housing that can withstand long-term storage and re-use.

Platform Approach
In addition to a full portfolio of sorbents, Pall Life Sciences is a leader in the drive towards continuous bioprocessing, introducing one of the industry’s first fully continuous downstream bioprocess solutions to the industry this year.

To learn more about building platform solutions with Pall Life Sciences portfolio of enabling products and services, please visit: www.pall.com/biopharm



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