Reversed-phase electrochromatography of amino acids and peptides using porous polymer monoliths |
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Authors: | Shediac R Ngola S M Throckmorton D J Anex D S Shepodd T J Singh A K |
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Affiliation: | BioSepra Process Chromatography, Life Technologies, A Division of Invitrogen Corporation, Rockville, MD 20850, USA. wschwart@lifetech.com |
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Abstract: | Efficient harvest and recovery of high-purity monoclonal antibodies was achieved using hydrophobic charge induction chromatography (HCIC). Both simple and complex feedstocks were studied, including protein-free cell culture supernatant and the clarified/concentrated milk of transgenic goats. Viral clearance studies demonstrated a 4-log reduction of MVM virus (minute virus of mice), along with substantial reduction of DNA content. Sorbent characterization studies confirmed that HCIC is based on the pH-dependent behavior of a dual-mode, ionizable ligand. Binding, based on hydrophobic interaction, was achieved under near-physiological conditions, and in the absence of lyotropic salt. Desorption was accomplished under mild conditions--pH 4.0. At this pH, both ligand and antibody carry a net positive charge, and desorption occurs on the basis of electrostatic charge repulsion. pH-based control of chromatographic function was demonstrated. Chromatography on this antibody-selective HCIC sorbent was evaluated as a cost-effective, process-compatible alternative to affinity chromatography protein A sorbents. |
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