Clinical-scale high-throughput human plasma proteome clinical analysis: Lung adenocarcinoma
Clinical-scale high-throughput human plasma proteome clinical analysis: Lung adenocarcinoma
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DOI：
10.1002/pmic.200401145
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发表时间：
2005-03-01
期刊：
影响因子：
3.4
通讯作者：
Nishimura, T
中科院分区：
文献类型：
--
作者：
Fujii, K;Nakano, T;Nishimura, T
临床蛋白质组学需要对大量人体样本进行稳定和可重复的分析。我们报告了一个高通量的全面的蛋白质分析系统，包括一个全自动的，在线的，二维微流液相色谱/串联质谱（2-D μ LC-MS/MS）系统用于临床蛋白质组学。线性离子阱质谱仪（ITMS）也被称为2-D ITMS仪器，其特征在于高扫描速度，被纳入μ LC-MS/MS系统，以获得高度提高的灵敏度和分辨率在MS/MS采集。该系统被用于评估牛血清白蛋白和人26 S蛋白酶体。这些高通量μ LC条件和2-D ITMS的应用导致蛋白质鉴定的灵敏度增加10倍。此外，从26 S蛋白酶体的肽片段被确定为三倍更有效地比传统的3-D ITMS仪器。在这项研究中，2-D μ LC-MS/MS系统，使用线性2-D ITMS已被应用于血浆蛋白质组分析的一些样本，从健康人和肺腺癌患者。使用2-D和1-D mu LC-MS/MS分析，在每个HSA和IgG耗尽的样品中分别检测到约250和100种不同的蛋白质，其对应于仅0.4 μ L的血浆。自动操作使得整个1-D和2-D mu LC-MS/MS分析的单次运行在11小时内完成。对从健康和腺癌组的蛋白质鉴定数据集提取的数据的研究表明，几种组特异性蛋白质可以是人血浆中表达的候选蛋白质疾病标志物。因此，它被证明，这种高通量μ LC-MS/MS蛋白质谱系统将实际上适用于蛋白质疾病标志物的发现，这是临床血浆蛋白质组项目的主要目标。
Clinical proteomics requires the stable and reproducible analysis of a large number of human samples. We report a high-throughput comprehensive protein profiling system comprising a fully automated, on-line, two-dimensional microflow liquid chromatography/tandem mass spectrometry (2-D mu LC-MS/MS) system for use in clinical proteomics. A linear ion-trap mass spectrometer (ITMS) also known as a 2-D ITMS instrument, which is characterized by high scan speed, was incorporated into the mu LC-MS/MS system in order to obtain highly improved sensitivity and resolution in MS/MS acquisition. This system was used to evaluate bovine serum albumin and human 26S proteasome. Application of these high-throughput mu LC conditions and the 2-D ITMS resulted in a 10-fold increase in sensitivity in protein identification. Additionally, peptide fragments from the 26S proteasome were identified three-fold more efficiently than by the conventional 3-D ITMS instrument. In this study, the 2-D mu LC-MS/MS system that uses linear 2-D ITMS has been applied for the plasma proteome analysis of a few samples from healthy individuals and lung adenocarcinoma patients. Using the 2-D and 1-D mu LC-MS/MS analyses, approximately 250 and 100 different proteins were detected, respectively, in each HSA- and IgG-depleted sample, which corresponds to only 0.4 mu L of blood plasma. Automatic operation enabled the completion of a single run of the entire 1-D and 2-D mu LC-MS/MS analyses within 11 h. Investigation of the data extracted from the protein identification datasets of both healthy and adenocarcinoma groups revealed that several of the group-specific proteins could be candidate protein disease markers expressed in the human blood plasma. Consequently, it was demonstrated that this high-throughput mu LC-MS/MS protein profiling system would be practically applicable to the discovery of protein disease markers, which is the primary objective in clinical plasma proteome projects.