Process optimisation for anion exchange monolithic chromatography of 4.2 kbp plasmid vaccine (pcDNA3F)

Clarence M. Ongkudon, and Michael K Danquah, (2010) Process optimisation for anion exchange monolithic chromatography of 4.2 kbp plasmid vaccine (pcDNA3F). Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences, 828 (28). pp. 2719-2725. ISSN 1570-0232

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Abstract

Anion exchange monolithic chromatography is increasingly becoming a prominent tool for plasmid DNA purification but no generic protocol is available to purify all types of plasmid DNA. In this work, we established a simple framework and used it to specifically purify a plasmid DNA model from a clarified alkaline-lysed plasmid-containing cell lysate. The framework involved optimising ligand functionalisation temperature (30–80 ◦C), mobile phase flow rate (0.1–1.8 mL/min), monolith pore size (done by changing the porogen content in the polymerisation reaction by 50–80%), buffer pH (6–10), ionic strength of binding buffer (0.3–0.7 M) and buffer gradient elution slope (1–10% buffer B/min). We concluded that preferential pcDNA3F adsorption and optimum resolution could be achieved within the tested conditions by loading the clarified cell lysate into 400 nm pore size of monolith in 0.7 M NaCl (pH 6) of binding buffer followed by increasing the NaCl concentration to 1.0 M at 3%B/min

Item Type: Article
Uncontrolled Keywords: Process optimisation, Anion exchange, Monolithic chromatography, Plasmid DNA pcDNA3F
Subjects: Q Science > QH Natural history
Divisions: SCHOOL > Biotechnology Research Institute (BRI)
Depositing User: Noraini
Date Deposited: 11 Mar 2019 00:24
Last Modified: 11 Mar 2019 00:24
URI: http://eprints.ums.edu.my/id/eprint/21552

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