Purification of a Large Protein Using Ion-Exchange Membranes

Abstract

Anion-exchange membranes were evaluated for the capture of a small protein (α-lactalbumin, 3.5-nm diameter) and a large protein (thyroglobulin, 20-nm diameter). The static binding capacity equaled the dynamic binding capacity and increased with increasing protein size. This result was in agreement with calculations based on monolayer coverage on the membrane surface and an absence of mass-transfer limitations. In contrast, for anion-exchange beads, the static capacity was the same for both proteins, and the dynamic capacity decreased strikingly with increasing protein size. These observations were attributed to very slow intrapore diffusion for large proteins in the beads, resulting in surface binding only. This work has important applications in the selection of chromatography media for the purification of viruses and plasmid DNA. Specifically, membranes with a high capacity for large biomolecules (20−300 nm) and a low capacity for small host-cell proteins and endotoxin contaminants are preferable to beads for the purification of such biomolecules.

Purification of a recombinant baculovirus of Autographa californica M nucleopolyhedrovirus by ion exchange membrane chromatography

“…In contrast to sucrose density centrifugation, aggregation of viral particles and loss of viral activity due to hydrodynamic shear stress are decreased by the use of ion exchange chromatography (Barsoum, 1999). Compared to conventional resin-based chromatography, IEMC shows relatively high capacities for biological macromolecules and a decreased sensitivity to flow rates affecting product yield and purity (Yang et al, 2002).…”

Process and economic evaluation for monoclonal antibody purification using a membrane‐only process

“…By switching to membrane-based adsorption/chromatography, the pressure drop is comparatively much lower across the membrane and the biomolecules can reach the target adsorption site without internal diffusion, thus allowing for higher flow rates. [22][23][24][25] However, the binding capacity of these membrane adsorbers has been reported to be significantly lower when compared with the binding capacities of similar resins. 26 To overcome the capacity limitation of membrane adsorbers, electrospun cellulose acetate nanofibrous membranes with nanofiber diameter of $500 nm have been developed.…”

Understanding the mechanism of virus removal by Q sepharose fast flow chromatography during the purification of CHO‐cell derived biotherapeutics

“…Ranging from 15 to 350 nm in diameter, viruses are quite large compared with most proteins. This large size leads to reduced diffusion coefficients for VP and other macromolecules and, combined with the relatively small pore sizes of most commercially available resins, limits most viruses to binding at the surface of the chromatography beads (Trilisky and Lenhoff, 2007;Yamamoto and Miyagawa, 1999;Yang et al, 2002;Yao and Lenhoff, 2006). Based on the unique nature of VP, it needs to be established empirically whether their behavior on anion exchange resins is consistent with the behaviors of proteins.…”