Protein diffusion in charged polyacrylamide gels. Visualization and analysis.

Protein diffusion in anionic, cross-linked polyacrylamide-based gels supported in fused-silica capillaries was characterized by a direct visualization method. Microphotography was used to obtain transient protein concentration profiles in these gels using cytochrome c as a probe molecule. Gels based on acrylamido-methylpropane sulfonic acid with 2.5-10% N,N'-methylene-bisacrylamide as a cross-linker and with a total polymer concentration of 0.21 g/cm3 yielded diffuse protein concentration profiles which were quantitatively consistent with a Fickian diffusion model. An analytical method was developed to calculate the diffusivity as a function of protein concentration in the gel from the experimental profiles. The diffusivity was found to assume values in the range 2.5-5.5x10(-8) cm2/s and varied somewhat with the protein concentration in the gel. The effects of some of the polymer properties, such as cross-link density, polymer concentration and charge, were also investigated for a limited range of conditions to derive qualitative trends. Results showed that the transport rates increased with a decrease in the cross-link density, were extremely reduced when the polymer concentration was doubled, and were slightly increased when the charge density was decreased by half by polymerizing a 1:1 mixture of acrylamide and acrylamido-methylpropane sulfonic acid monomers.