Foucher Aude L, Craft David R, Gelfand Craig A
BD Diagnostic, Franklin Lakes, NJ, USA.
Methods Mol Biol. 2010;641:27-45. doi: 10.1007/978-1-60761-711-2_3.
Urine is a complex fluid, which is thought to contain valuable diagnostic information regarding general health. In particular, there is great diagnostic potential in the peptide and/or protein content of urine, but the information is present in low abundance. Most traditional proteomic techniques lack sufficient sensitivity/dynamic range, especially for dilute and/or complex samples. However, orthogonal separation methods can be applied prior to protein/peptide analysis to increase the success rate of urine proteomic studies and access this potentially valuable information. In this chapter, we describe isoelectric focusing (IEF) of intact urine proteins, via free flow electrophoresis (FFE), prior to typical peptide-based mass spectrometry analysis, facilitating the deep analysis of urine protein detection and identification, for biomarker discovery. Our work demonstrates that such an approach can be used as a preprocessing step and can be integrated into a workflow for the successful identification of protein components (biomarkers) from urine.
尿液是一种复杂的液体，被认为含有有关总体健康状况的宝贵诊断信息。特别是，尿液中的肽和/或蛋白质含量具有很大的诊断潜力，但这些信息的丰度较低。大多数传统蛋白质组学技术缺乏足够的灵敏度/动态范围，尤其是对于稀释和/或复杂的样本。然而，在进行蛋白质/肽分析之前，可以应用正交分离方法来提高尿液蛋白质组学研究的成功率，并获取这些潜在的有价值信息。在本章中，我们描述了在基于肽的典型质谱分析之前，通过自由流动电泳（FFE）对完整尿液蛋白质进行等电聚焦（IEF），以促进尿液蛋白质检测和鉴定的深度分析，用于生物标志物发现。我们的工作表明，这种方法可以用作预处理步骤，并可以整合到工作流程中，以成功鉴定尿液中的蛋白质成分（生物标志物）。
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