Preparation of liposome membrane adsorbers and testing for plasmid purification
•Hydrophobic matrices were prepared from pre-activated commercial membrane adsorbers.•Liposomes were bound to pre-chromatographic membranes (7 ± 2 phospholipids μmol/cm2).•CLSM of functionalized membrane shows fibrous structure and adsorbed plasmid.•Plasmid DNA from cell lysates was bound and eluted in membrane chromatography (HIC).•Liposome-membrane chromatographic performances are assessed and discussed.
DNA therapeutics can prevent or overcome several diseases and use plasmid DNA (pDNA) to deliver specific genes to the hosts for the therapies. The demand for highly purified pDNA is increased. Hydrophobic interaction chromatography (HIC) using membrane adsorbers is a purification technique able to separate lysate contaminants from pDNA, with improved productivity when compared to the use of classic adsorbers but still relying on hydrophobic interactions between patches on the surface of biomolecules and those designed at the chromatographic matrix.A final step in the downstream processing of pDNA was performed by HIC on membrane adsorbers functionalized with unilamellar liposomes of DOPE, DOPC, DLPA and DSPA. The chemical and morphological fibre structure characterizations of the derived membrane adsorbers are disclosed.Confocal microscope images demonstrated the ability of these membranes to interact with pDNA under high kosmotropic salt concentration environment. The study of chromatographic performance of the adsorbers on pDNA purification showed that DLPA-membrane has acquired the lowest ligand density but a considerable membrane adsorbing capacity; conversely, the DOPC-membrane has highest binding capacity and high HIC performance parameters. Purification yields obtained are still lower than those found for other chromatographic membrane adsorbers, consistent with the promotion of unspecific interactions hindering pDNA purification. This work gives a new insight onto liposome-based membrane adsorbers application for the pDNA downstream purification process.
Journal: Biochemical Engineering Journal - Volume 93, 15 January 2015, Pages 1–10