Characterization of a solvent system for separation of water-insoluble poliovirus proteins by reversed-phase high-performance liquid chromatography.

Formic acid in high concentration is an extremely potent solvent for proteins, particularly for hydrophobic ones. 60% Formic acid, necessary for solubilization of structural polypeptides of poliovirus and other proteins, modified at the cysteines, was used together with 2-propanol or acetonitrile as organic modifier for gradient elution in reversed-phase high-performance liquid chromatography. Several reversed-phase columns were tested. In each case, polypeptides were eluted quantitatively. It was demonstrated that this solvent system, with its high proportion of formic acid, did not affect the size, hydrophobicity and charge of the separated polypeptides. By injection into rabbits of poliovirus polypeptides, obtained in high purity by chromatography in the new solvent system, monospecific antibodies were induced, the specificity of which was determined by immunoprecipitation.