Adaptation of pulsed-field gel electrophoresis for the improved fractionation of spheres

Philip Serwer, Gary A. Griess

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

During gel electrophoresis, proteins are fractionated by both size (radius, R, for a sphere) and average electrical surface charge density. The fractionation by size depends entirely on nonspecific steric and hydrodynamic effects (sieving) of the fibrous network that forms the gel. Previous studies have demonstrated the following aspects of sieving: (1) as a function of the radius of the effective pore of a gel (P(E)), electrophoretic mobility decreases in magnitude by a factor that has the form 1-a1(R/P(E))+a2(R/P(E))(n); a1 and a2 are constants; a1 is always positive; a2 is usually positive. (2) For a nonspherical particle not preferentially oriented in the gel, the effective R is best approximated by the R of a sphere that has a surface area equal to that of the nonspherical particle. For spheres, resolution by R is progressively increased by lowering P(E), but maintaining P(E)>R; this approach, however, progressively increases the time of electrophoresis. To improve fractionation of both proteins and multimolecular protein complexes, a ratcheting-based procedure of pulsed-field gel electrophoresis is introduced here for spherical particles. Successfully tested here with latex spheres 95-235nm in R, this procedure bypasses limitations of conventional sieving-based separations. Even though all spheres are negatively charged, larger spheres can be made to migrate in a direction opposite to the direction in which smaller spheres migrate. Copyright (C) 1998 Elsevier Science B.V.

Original languageEnglish (US)
Pages (from-to)299-306
Number of pages8
JournalAnalytica Chimica Acta
Volume372
Issue number1-2
DOIs
StatePublished - Oct 19 1998

Keywords

  • Fractionation
  • Gel, sieving by
  • Gel, structure of
  • Gel, trapping by
  • Latex sphere
  • Protein

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Environmental Chemistry
  • Spectroscopy

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