Templated pores in hydrogels for improved size selectivity in gel permeation chromatography.

The pore structures of cross-linked polyacrylamide gels can be altered by polymerizing in the presence of high concentrations of unreactive, micellar surfactant cosolutes which act as "templates". Removal of surfactant after polymerization is expected to leave pores with the approximate shape and dimensions of the surfactant micelles. A simple model was developed to simulate gel permeation chromatography (GPC) separations of globular proteins on templated gels. The model assumes that the partition coefficient for sieving of a protein is equal to the fraction of gel volume accessible to a sphere with a radius equal to the protein Stokes radius. The total gel volume is considered to include a fraction that is a conventional, random gel matrix and a remaining fraction contributed by templated pores. The pore size distribution of the conventional gel was estimated using the Ogston equation, which approximates the matrix as a random collection of long, thin, rigid fibers. Templated pores were assumed to have a Gaussian distribution of radii centered about some mean determined by the micelle radius. In comparison to conventional media, gels with templated pores are predicted to exhibit more sharply defined exclusion limits and improved resolution over a narrow size range centered on the mean templated pore size. Selectivity and resolution are expected to increase as the volume fraction of templated pores is increased and as the dispersion of templated pore radius is decreased. Small changes in template radius lead to large changes in the molecular weight range of optimal separation of globular proteins. It should be possible to create a series of GPC media that collectively offer high resolution over the molecular weight range of most globular proteins of interest.