Author/Authors :
Davis، نويسنده , , Michael T. and Beierle، نويسنده , , Jill and Bures، نويسنده , , Edward T. and McGinley، نويسنده , , Michael D. and Mort، نويسنده , , Jessica and Robinson، نويسنده , , John H. and Spahr، نويسنده , , Chris S. and Yu، نويسنده , , Wen and Luethy، نويسنده , , Roland and Patterson، نويسنده , , Scott D.، نويسنده ,
Abstract :
A simple multidimensional liquid chromatography system utilizing an isocratic pump and a HPLC system is described for the comprehensive proteomic analysis of complex peptide digest mixtures by coupled LC–LC–MS–MS techniques. A binary ion-exchange separation was achieved through the use of a strong cation-exchange column followed by a reversed-phase column for data-dependent LC–MS–MS analysis of the unbound analytes, and following salt elution (and concomitant column reequilibration), the bound analytes. Off-line validation of the platform showed near quantitative recovery of fractionated peptides and essentially complete ion-exchange partitioning. In comparative analyses of a highly complex peptide digest mixture a >40% increase in the number of peptide and protein identifications was achieved using this multidimensional platform compared to an unfractionated control.
