Analytical Biochemistry, Department of Pharmacy, University of Groningen, Groningen, The Netherlands.
J Sep Sci. 2010 Jun;33(10):1421-37. doi: 10.1002/jssc.201000050.
Multidimensional chromatography coupled to mass spectrometry (LC(n)-MS) provides more separation power and an extended measured dynamic concentration range to analyse complex proteomics samples than one dimensional liquid chromatography coupled to mass spectrometry (1D-LC-MS). This review gives an overview of the most important aspects of LC(n)-MS with respect to optimizing peak capacity and evaluate orthogonality. We review recent developments in LC(n)-MS to analyse proteomics samples from the analyst point of view and give an overview over methods and future developments to process LC(n)-MS data for comprehensive differential protein expression profiling. Examples from our research, such as combining protein fractionation using high temperature reverse phase (RP) columns followed by analysis of the trypsin-digested fractions by RP LC-MS, serve to highlight possibilities and shortcomings of present-day approaches. Other LC(n)-MS systems that have been used to analyse highly complex shotgun proteomic samples, such as the combination of RP columns using low and high pH eluents or the combination of hydrophilic interaction liquid chromatography (HILIC) with RP-MS is discussed in detail.
多维色谱与质谱联用（LC(n)-MS）比一维液相色谱与质谱联用（1D-LC-MS）提供了更多的分离能力和扩展的测量动态浓度范围，可用于分析复杂的蛋白质组学样品。本文综述了 LC(n)-MS 的最重要方面，涉及如何优化峰容量和评估正交性。我们从分析人员的角度综述了 LC(n)-MS 分析蛋白质组学样品的最新进展，并概述了用于全面差异蛋白质表达分析的 LC(n)-MS 数据处理方法和未来发展。我们的研究示例，如使用高温反相（RP）柱进行蛋白质分级，然后用 RP LC-MS 分析胰蛋白酶消化的级分，突出了当前方法的可能性和局限性。还详细讨论了其他用于分析高度复杂的鸟枪法蛋白质组学样品的 LC(n)-MS 系统，如使用低 pH 和高 pH 洗脱剂的 RP 柱的组合，或亲水相互作用液相色谱（HILIC）与 RP-MS 的组合。
