Impact of grafting on the design of new membrane adsorbers for adenovirus purification

• Hydrogel grafted membranes are more suitable for virus purification than direct grafted ones.
• Virus recovery is better for hydrogel grafted ligand densities of 1.6–2.7 μmol/cm2 and increased NaCl in the feed.
• Direct grafting requires ligand densities >4.5 μmol/cm2 and NaCl concentrations <25 mM for improved virus recovery.
• DNA's 10%-DBC increases 5-fold when the ligand density is increased from 1.7 to 3.3 μmol/cm2; this is not seen for virus.
• A novel 96-well plate system coupled with a small-scale device minimizes the amount of bulk material used for this study.

The impacts of quaternary amine ligand density and matrix structure, namely hydrogel grafted and directly grafted, on state-of-the-art chromatographic membranes operated in bind-and-elute mode were evaluated for the purification of adenovirus serotype 5. The experiments were performed on a 96-well plate membrane holder, which is a convenient high-throughput screening tool for obtaining the best operating conditions for a process yield optimization. The results show that the hydrogel-grafted membranes are more suitable for virus purification than the directly grafted ones. By reducing the number of grafted ligands to low (1.7 μmol/cm2) or medium (2.4 μmol/cm2) density, it is possible to increase the recovery of purified virus by 60% compared to a highly charged membrane (3.3 μmol/cm2) that yielded a recovery rate lower than 30%. In the reported experiments, Sartobind® Q, chosen as benchmark comparison, provides a better compromise between high recovery and large dynamic binding capacity. Overall, this work contributes to the understanding and development of new membrane adsorbers specifically designed for virus purification.