title: Protein purification by Off-Gel electrophoresis.
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 (pI) 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 pI 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 coli extract is shown to illustrate the possible applications.
authors.affiliation: Laboratoire d'Electrochimie, Département de Chimie, Ecole Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Ros
authors.forename: Michel
authors.initials: M
authors.lastname: Faupel
authors.forename: Hervé
authors.initials: H
authors.lastname: Mees
authors.forename: Jan van
authors.initials: Jv
authors.lastname: Oostrum
authors.forename: Rosaria
authors.initials: R
authors.lastname: Ferrigno
authors.forename: Frédéric
authors.initials: F
authors.lastname: Reymond
authors.forename: Philippe
authors.initials: P
authors.lastname: Michel
authors.forename: Joël S
authors.initials: JS
authors.lastname: Rossier
authors.forename: Hubert H
authors.initials: HH
authors.lastname: Girault
mesh_terms: D001426:Bacterial Proteins / Q000302:isolation & purification; D015180:Electrophoresis, Gel, Two-Dimensional / Q000295:instrumentation / Q000379:methods*; D004926:Escherichia coli / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D011506:Proteins / Q000302:isolation & purification*; D020543:Proteome / Q000302:isolation & purification; D015843:Serpins / Q000302:isolation & purification
chemical_list: D001426:Bacterial Proteins; D011506:Proteins; D020543:Proteome; D015843:Serpins