Partitioning in aqueous two-phase systems: Analysis of strengths, weaknesses, opportunities and threats.

For half a century aqueous two-phase systems (ATPSs) have been applied for the extraction and purification of biomolecules. In spite of their simplicity, selectivity, and relatively low cost they have not been significantly employed for industrial scale bioprocessing. Recently their ability to be readily scaled and interface easily in single-use, flexible biomanufacturing has led to industrial re-evaluation of ATPSs.

Purification of pegylated proteins.

Separation of PEGylated proteins is challenging because PEG itself is a relatively inert, neutral, hydrophilic polymer and the starting point for PEGylation is a pure protein. Thus, other than molecular weight and size, differences in the physicochemical properties typically used to fractionate proteins may be slight between different PEGylated forms of a protein. The usual properties of electrostatic charge and molecular weight (size) form the basis of the most commonly used separation techniques, particularly IEC, SEC, and ultrafiltration.

Novel in situ polymerized coatings for hydrophobic interaction chromatography media.

Hydrophobic interaction chromatography (HIC) and other capture media are typically produced by grafting different ligands to base matrices at defined surface densities. This often complicates media production. An alternative approach to media involving in situ radical initiated polymerization was used to graft polymer coatings directly at Sepharose(R) polymeric base matrices.

Ion exchange chromatography of antibody fragments.

Effects of pH and conductivity on the ion exchange chromatographic purification of an antigen-binding antibody fragment (Fab) of pI 8.0 were investigated. Normal sulfopropyl (SP) group modified agarose particles (SP Sepharosetrade mark Fast Flow) and dextran modified particles (SP Sepharose XL) were studied.

Modeling of protein interactions with surface-grafted charged polymers. Correlations between statistical molecular modeling and a mean field approach.

Ion exchange media involving charge groups attached to flexible polymers are widely used for protein purification. Such media often provide enhanced target protein purity and yield. Yet, little is understood about protein interaction with such media at the molecular level, or how different media architectures might affect separation performance.

Prediction of the viscosity radius and the size exclusion chromatography behavior of PEGylated proteins.

Size exclusion chromatography (SEC) was used to determine the viscosity radii of equivalent spheres for proteins covalently grafted with poly(ethylene glycol) (PEG). The viscosity radius of such PEGylated proteins was found to depend on the molecular weight of the native protein and the total weight of grafted PEG but not on PEG molecular weight, or PEG-to-protein molar grafting ratio. Results suggest grafted PEG's form a dynamic layer over the surface of proteins.