The sieving of spheres during agarose gel electrophoresis: Quantitation and modeling

✍ Scribed by Gary A. Griess; Elena T. Moreno; Richard A. Easom; Philip Serwer

• Publisher: Wiley (John Wiley & Sons)
• Year: 1989
• Tongue: English
• Weight: 590 KB
• Volume: 28
• Category: Article
• ISSN: 0006-3525
• DOI: 10.1002/bip.360280811

✦ Synopsis

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. Nutl. Acud. Scc. USA 65,970-9771 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-o.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.