LC–tandem MS in proteome characterization

Abstract

The coupling of high-performance liquid chromatography (HPLC) to mass spectrometry (MS) has been routinely used for a number of years for the analysis of a wide variety of different compounds. In typical proteomic analyses, where enzymatic digestion is used to generate proteolytic peptides, the limited amount of sample restricts the utility of conventional HPLC methods for MS detection. As reduced column diameters and nanolitre per minute flow rates have become increasingly standard, the application of HPLC to the analysis of low-volume, low-abundance samples has now become readily achievable. A number of novel chromatographic methods have increased the utility of such approaches for proteome-wide analyses. However, there remain in proteomic analyses some important challenges, which are being addressed by state-of-the-art methodologies. This article reviews a number of pertinent considerations and technological advances in proteomic analyses using HPLC–tandem MS (MS 2).