Cation exchange chromatography in antibody purification: pH screening for optimised binding and HCP removal

Stein, Andreas; Kiesewetter, André

doi:10.1016/j.jchromb.2006.10.010

Cited by 72 publications

(28 citation statements)

References 18 publications

(22 reference statements)

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“…We have utilized 1 mL columns as opposed to plate-based screening (Stein and Kiesewetter, 2007) for evaluation of the resins to enable rapid scale-up of the process and to measure the impact of residence time in a packed bed (as opposed to determining the equilibrium performance that is obtained from plate-or batch-mode experiments). Some disadvantages of this scale of operation that can impact the resolution of impurities are wall effects and small plate numbers, owing to the short bed height.…”

Section: Introductionmentioning

confidence: 99%

Maximizing productivity of chromatography steps for purification of monoclonal antibodies

Tugçu

Roush

Göklen

2007

Biotech & Bioengineering

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The large scale production of monoclonal antibodies presents a challenge to design efficient and cost effective downstream purification processes. We explored a two stage resin screening approach to identify the best candidates to be utilized for the platform purification of monoclonal antibodies. The study focused on commercially available affinity resins including Protein A, mimetic and mixed-mode interaction resins as well as ion exchangers used in polishing steps. An initial screening using pure proteins was followed by a final screening where selected resins were utilized for the purification of MAbs in complex mixtures. Initial screenings aimed to measure the theoretical upper limit for dynamic binding capacity (DBC) at 1% breakthrough and productivity. We confirmed that DBC of affinity, mimetic and mixed-mode resins was a strong function of the linear velocity used for loading. Productivities >27 g/(L-h), were obtained for rProtein A FF, Mabselect and Prosep rA Ultra at 2 min residence time. For the cation exchangers, we identified UNOsphere S and Fractogel SO(3) as the best candidates for our purification based on DBC. For anion exchangers operated in flowthrough mode, Q Sepharose XL and UNOsphere Q were selected from the initial screening based on DBC and resolution of IgG from BSA. Finally, a three step purification scheme was implemented using the selected affinity and ion exchangers for the purification of IgG from complex feedstocks. We found that Mabselect followed by UNOsphere Q and UNOsphere S provided the best purification scheme for our applications based on productivity.

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Section: Introductionmentioning

confidence: 99%

Maximizing productivity of chromatography steps for purification of monoclonal antibodies

Tugçu

Roush

Göklen

2007

Biotech & Bioengineering

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“…However, adsorbents incorporating Protein A, as well as Protein G from Streptococcus sp. , do not bind to certain mammalian antibody isotypes, show sensitivity to alkaline conditions during clean‐in‐place (CIP) procedures, and exhibit incomplete clearance of host cell proteins, lipids and nucleic acids (Andrew and Titus, ; Huse et al , ; Ishihara et al , ; Stein and Kiesewetter, ; Samaranayake et al , ; Sisodiya et al , ; Zhang et al , ). As a consequence, a variety of research strategies have been explored during the past decade with the aim to develop suitable substitutes for these proteinaceous ligands, and in this manner address these downstream processing constraints.…”

Section: Introductionmentioning

confidence: 99%

Studies on the binding sites of IgG2 monoclonal antibodies recognized by terpyridine‐based affinity ligands

Lin

Boysen

Campi

et al. 2016

J of Molecular Recognition

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This investigation has examined the origin of the molecular recognition associated with the interaction of monoclonal IgG2's with terpyridine-based ligands immobilized onto agarose-derived chromatographic adsorbents. Isothermal titration calorimetric (ITC) methods have been employed to acquire thermodynamic data associated with the IgG2-ligand binding. These ITC investigations have documented that different enthalpic and entropic processes are involved depending on the nature of the chemical substituents in the core structure of the terpyridinyl moiety. In addition, molecular docking studies have been carried out with IgG2 structures with the objective to identify possible ligand binding sites and key interacting amino acid residues. These molecular docking experiments with the different terpyridine-based ligands have shown that all of the examined ligands can potentially undergo favorable interactions with a site located within the Fab region of the IgG2. However, another favorable binding site was also identified from the docking poses to exist within the Fc region of the IgG2 for some, but not all, of the ligands studied. These investigations have provided a basis to elucidate the unique binding properties and chromatographic behaviors shown by several substituted terpyridine ligands in their interaction with IgGs of different isotype. Copyright © 2016 John Wiley & Sons, Ltd.

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“…In cation exchange chromatography, positively charged amino acid side chains of the protein molecule interact with the negatively charged ligands of the chromatography matrix. [6,7] Ion exchange column chromatography has been widely used for purification of biotherapeutic proteins because of its high capacity and high efficiency. Most commercial available ion exchange beads are based on sephadex, agarose, silica and cross linked cellulose.…”

Section: Introductionmentioning

confidence: 99%

Use of polymeric membranes for purification of an E. coli expressed biotherapeutic protein

Muthukumar

Rathore

2015

Preparative Biochemistry & Biotechnology

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Polymers have had a significant impact on the field of bioseparations in the past few decades. Most recently, membrane chromatography has emerged as an efficient alternative to the conventional packed-bed chromatography by eliminating the diffusion-related limitations associated with the traditional resin beads. In this article, we examine six membrane adsorbers for purification of granulocyte colony-stimulating factor (GCSF), an Escherichia coli-based biotherapeutic. These adsorbers differ either in their base matrix or in the surface chemistry. The role of interactions between the filter surfaces and the protein molecules in effecting these separations is the focus of the article.

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Cation exchange chromatography in antibody purification: pH screening for optimised binding and HCP removal

Cited by 72 publications

References 18 publications

Maximizing productivity of chromatography steps for purification of monoclonal antibodies

Maximizing productivity of chromatography steps for purification of monoclonal antibodies

Studies on the binding sites of IgG2 monoclonal antibodies recognized by terpyridine‐based affinity ligands

Use of polymeric membranes for purification of an E. coli expressed biotherapeutic protein

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