Free-flow electrophoresis of the human urinary proteome.
Prefractionation of complex protein samples prior to mass spectrometry provides a method for the isolation of low-abundance proteins into specific fractions thereby enabling their identification. Free-flow electrophoresis in the isoelectric focusing mode (IEF-FFE) presents a complementary approach to established prefractionation methodologies. Proteins are separated in solution according to their isoelectric point (pI) with a high throughput of sample volume. The separation may be performed under denaturing or nondenaturing conditions and detergents may be added to promote protein solubilization. A protocol covering the pH range from pH 3 to 9 under denaturing conditions was used to illustrate the method of IEF-FFE including sample preparation prior to reversed-phase liquid chromatography and tandem mass spectrometry. The IEF-FFE separation was applied to a sample of human urine.
Nissum
Mikkel
M
BD Diagnostics, Martinsried, Germany.
Wildgruber
Robert
R
D004586:Electrophoresis / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D007525:Isoelectric Focusing / Q000295:instrumentation / Q000379:methods; D013058:Mass Spectrometry / Q000295:instrumentation / Q000379:methods; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000295:instrumentation / Q000379:methods*; D014556:Urine / Q000737:chemistry*
D020543:Proteome