Diffusion of macromolecules in agarose gels: Comparison of linear and globular configurations

The diffusion coefficients (D) of different types of macromolecules (protei ns, dextrans, polymer beads, and DNA) were measured by fluorescence recover y after photobleaching (FRAP) both in solution and in 2% agarose gels to co mpare transport properties of these macromolecules. Diffusion measurements were conducted with concentrations low enough to avoid macromolecular inter actions. For gel measurements, diffusion data were fitted according to diff erent theories: polymer chains and spherical macromolecules were analyzed s eparately. As chain length increases, diffusion coefficients of DNA show a clear shift from a Rouse-like behavior (D-G congruent to N-0(-0.5)) to a re ptational behavior (D-G similar or equal to N-0(-2.0)). Th, pore size, a, o f a 2% agarose gel cast in a 0.1 M PBS solution was estimated. Diffusion co efficients of the proteins and the polymer beads were analyzed with the Ogs ton model and the effective medium model permitting the estimation of an ag arose gel fiber radius and hydraulic permeability of the gels. Not only did flexible macromolecules exhibit greater mobility in the gel than did compa rable-size rigid spherical particles, they also proved to be a more useful probe of available space between fibers.