Screening of effective column rinse solvent for Protein-A chromatography

Abstract

Mildly acidic arginine solution is highly effective in elution of bound proteins from Protein-A columns. Although Protein-A is specific in antibody capture, it does bind other proteins, which must then be removed before elution by aqueous arginine solution. If they are not removed, a strong elution property of aqueous arginine solutions will elute the contaminating proteins along with antibodies. Here we have examined various salt solutions as a column rinse solvent. We screened various solvents for their effects on binding of purified antibodies to Protein-A, instead of their effectiveness to elute the bound contaminants. Those solvents that result in a slight flow-through of the antibodies during loading should be effective in eluting non-specifically bound proteins that have weaker affinity for Protein-A than antibodies: namely, if a particular solvent reduces antibody binding to Protein-A, it is expected to be effective in reducing binding of contaminants and hence eluting them. Such screening showed a few compounds, including arginine and sodium acetate, as potential column rinse agents. A combination of arginine and sodium acetate was tested for a few crude materials containing antibodies.