Protein Transport in Constrained Anionic Hydrogels: Diffusion and Boundary-Layer Mass Transfer

The partitioning and transport of proteins in anionic, polyacrylamide-based gels were characterized by a direct visualization method using cytochrome c as the probe molecule. Homogeneous gels stabilized against mechanical and osmotic forces were made by synthesizing them inside fused-silica capillaries with a square cross section. Mixtures of 2-acrylamido-2-methylpropane sulfonic acid and acrylamide as the starting co-monomers and N, N ‘-methylene bisacrylamide as a cross-linker were used in preparing these gels. After polymerization and equilibration of the samples in a buffer, the diffusional transport of cytochrome c was studied via the microscopic determination of the evolution of concentration profiles in the gel upon exposure to a protein solution. Calibrated digitized profiles were used to determine the protein diffusivity in the gel and the effects of boundary-layer mass-transfer resistance on transient adsorption and desorption. The effects of protein and salt concentrations in solution and the effects of the gel composition were determined. The ranges of experimental conditions covered gels with cross-link densities between 2.5 and 10%, total polymer concentrations between 0.05 and 0.42 g/cm 3, and charge densities between 97 and 970 μequiv/cm 3 . In each case, the concentration profiles observed had a diffuse character and evolved in time in a manner consistent with Fick's law. However, the experimental diffusivity values varied with protein concentration in the gel and were strongly influenced by the composition of the gel and by the salt concentration in solution.