Label-free quantitative proteomics using large peptide data sets generated by nanoflow liquid chromatography and mass spectrometry
Label-free quantitative proteomics using large peptide data sets generated by nanoflow liquid chromatography and mass spectrometry
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DOI：
10.1074/mcp.t500039-mcp200
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发表时间：
2006-07-01
影响因子：
7
通讯作者：
Yamada, Tesshi
中科院分区：
文献类型：
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作者：
Ono, Masaya;Shitashige, Miki;Yamada, Tesshi
我们开发了一个由机器和软件模块组成的集成平台，可以将纳米流LC-MS生成的大量数据应用于差异蛋白质表达分析。未标记的蛋白质样品用改性胰蛋白酶完全消化，并通过低速（200 nl/min）一维HPLC分离。通过使用混合Q-TOF质谱仪的测量模式每1秒获得质谱，并显示在二维平面中，其中m/z值沿x轴沿着，保留时间沿y轴沿着显示。使用基于动态规划算法的新开发的软件来调整纳米LC的时间抖动。全面性（在60 μ g细胞蛋白样品中可检出高于预定阈值的60，000 - 160，000个峰），重现性（平均变异系数为0.35-0.39，重复间相关系数大于0.92），该平台的宽动态范围（超过103）的精确定量保证了其在各种类型的实验和翻译蛋白质组学中的应用。
We developed an integrated platform consisting of machinery and software modules that can apply vast amounts of data generated by nanoflow LC-MS to differential protein expression analyses. Unlabeled protein samples were completely digested with modified trypsin and separated by low speed (200 nl/min) one-dimensional HPLC. Mass spectra were obtained every 1 s by using the survey mode of a hybrid Q-TOF mass spectrometer and displayed in a two-dimensional plane with m/z values along the x axis, and retention time was displayed along the y axis. The time jitter of nano-LC was adjusted using newly developed software based on a dynamic programming algorithm. The comprehensiveness (60,000-160,000 peaks above the predetermined threshold detectable in 60-mu g cell protein samples), reproducibility (average coefficient of variance of 0.35-0.39 and correlation coefficient of over 0.92 between duplicates), and accurate quantification with a wide dynamic range (over 103) of our platform warrant its application to various types of experimental and translational proteomics.