Kim Yeoun Jin, Zhan Ping, Feild Brian, Ruben Steven M, He Tao
Celera, 45 West Gude Drive, Rockville, Maryland 20850, USA.
Anal Chem. 2007 Aug 1;79(15):5651-8. doi: 10.1021/ac070200u. Epub 2007 Jun 20.
The reproducibility of a given method for relative quantitation governs the reliability of liquid chromatography-mass spectrometry (LC-MS) based differential analysis in proteomic studies. Understanding the noise level introduced from biological, chemical, and instrumental sources not only helps to determine the experimental design but also aids in assessing the reliability of expression ratios used for quantitation. Here we present a reproducibility assessment method for relative quantitation based on the intensity ratio distribution of common features in LC-MS replicates. This method applies to both decoupled (label-free quantitation) and coupled (label-dependent quantitation) methods. Aligning the features of LC-MS maps directly for the decoupled method or by matching an LC-MS map and its virtual map for the coupled method results in a list of common features for replicate samples. We find that the ratio distribution of the common features successfully indicates the reproducibility of each experiment prior to MS/MS peptide sequencing in three different quantitation strategies: decoupled, coupled isotope-coded affinity tag, and coupled stable isotope labeling of amino acids in cell culture experiments.
在蛋白质组学研究中，特定相对定量方法的可重复性决定了基于液相色谱 - 质谱联用（LC-MS）的差异分析的可靠性。了解生物、化学和仪器来源引入的噪声水平，不仅有助于确定实验设计，还有助于评估用于定量的表达比率的可靠性。在此，我们基于LC-MS重复样品中共有特征的强度比分布，提出一种相对定量的可重复性评估方法。该方法适用于解耦（无标记定量）和耦合（标记依赖定量）方法。对于解耦方法，直接对齐LC-MS图谱的特征；对于耦合方法，通过匹配LC-MS图谱及其虚拟图谱，得到重复样品的共有特征列表。我们发现，在三种不同的定量策略中，即解耦、耦合同位素编码亲和标签以及细胞培养实验中氨基酸的耦合稳定同位素标记，共有特征的比率分布在MS/MS肽测序之前成功地表明了每个实验的可重复性。
