Protein purification by off-gel electrophoresis

Alexandra Ros, Michel Faupel, Hervé Mees, Jan Van Oostrum, Rosaria Ferrigno, Frédéric Reymond, Philippe Michel, Joël S. Rossier, Hubert H. Girault

Research output: Contribution to journalArticle

96 Scopus citations

Abstract

A novel free-flow protein purification technique based on isoelectric electrophoresis is presented, where the proteins are purified in solution without the need of carrier ampholytes. The gist of the method is to flow protein solutions under an immobilised pH gradient gel (IPG) through which an electric field is applied perpendicular to the direction of the flow. Due to the buffering capacity of the IPG gel, proteins with an isoelectric point (p/) close to pH of the gel in contact with the flow chamber stay in solution because they are neutral and therefore not extracted by the electric field. Other proteins will be charged when approaching the IPG gel and are extracted into the gel by the electric field. Both a demonstration experiment with p/ markers and a simulation of the electric field distribution are presented to highlight the principle of the system. In addition, an isoelectric fractionation of an Escherichia coil extract is shown to illustrate the possible applications.

Original languageEnglish (US)
Pages (from-to)151-156
Number of pages6
JournalProteomics
Volume2
Issue number2
DOIs
StatePublished - Jan 1 2002

Keywords

  • Isoelectric focusing
  • Isoelectric purification
  • Prefractionation

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

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    Ros, A., Faupel, M., Mees, H., Van Oostrum, J., Ferrigno, R., Reymond, F., Michel, P., Rossier, J. S., & Girault, H. H. (2002). Protein purification by off-gel electrophoresis. Proteomics, 2(2), 151-156. https://doi.org/10.1002/1615-9861(200202)2:2<151::AID-PROT151>3.0.CO;2-9