The sieving of spheres during agarose gel electrophoresis: quantitation and modeling.
By use of agarose gel electrophoresis, the sieving of spherical particles in agarose gels has been quantitated and modeled for spheres with a radius (R) between 13.3 and 149 nm. For quantitation, the electrophoretic mobility has been determined as a function of agarose percentage (A). Because a previously used model of sieving [D. Rodbard and A. Chrambach (1970) Proc. Natl. Acad. Sci. USA 65, 970-977] was found incompatible with some of these data, alternative models have been tested. By use of an underivatized agarose, two models, both based on the assumption of a single effective pore radius (PE) for each A, were found to yield PE values that were independent of R and that were in agreement with values of PE obtained independently (PE = 118 nm X A-0.74): sieving by altered hydrodynamics in a cylindrical tube of radius, PE, and sieving by steric exclusion from a circular hole of radius, PE. The same analysis applied to a 6.5% hydroxyethylated commercial agarose yielded a steeper PE vs A plot and also agreement of the above two models with the data. The PE vs A plot was significantly altered by both further hydroxyethylation and factors that cause variation in the electro-osmosis found in commercial agarose.
Griess
G A
GA
Moreno
E T
ET
Easom
R A
RA
Serwer
P
P
D001704:Biopolymers; D004586:Electrophoresis / Q000379:methods*; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D008962:Models, Theoretical; D008968:Molecular Conformation
D001704:Biopolymers