Simplifying Analysis of Biomolecules using Two-Dimensional LC-MS

Introduction :

Standard reversed-phase LC–MS and LC–MS–MS techniques have limited effectiveness in the analysis of complex biomolecule mixtures because of co-eluting species compromising MS–MS data collection. Components present in relatively small concentrations, such as small peptide fragments, can be neglected by the detection method or interpreted as background noise. Two-dimensional (2D) LC–MS methods increase the efficiency of protein identification as co-eluting components are separated into different fractions, in effect removing interferences from more abundant co-eluting species. This application note compares the data obtained when using a traditional one-dimensional method with reversed-phase chromatography using a silica-based C18 column, versus a 2D technique involving initial fractionation of peptides on the basis of their charge using a cation exchange column, followed by reversed-phase chromatography of each fraction.

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