Purification of a large protein using ion-exchange membranes

Anion-exchange membranes were evaluated for the capture of a small protein (a-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.