Robust Microflow LC-MS/MS for Proteome Analysis: 38 000 Runs and Counting
Robust Microflow LC-MS/MS for Proteome Analysis: 38 000 Runs and Counting
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DOI：
10.1021/acs.analchem.1c00257
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发表时间：
2021-02-17
影响因子：
7.4
通讯作者：
Kuster, Bernhard
中科院分区：
文献类型：
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作者：
Bian, Yangyang;Bayer, Florian P.;Kuster, Bernhard
微流液相色谱串联质谱（mu LC-MS/MS）正在成为一个可行的替代纳流LC-MS/MS的蛋白质组分析。我们最近已经证明了这种系统在1 mm内径下工作的潜力。X 150 mm柱，流速为50 μ L/ min，适用于高通量应用。在分析了过去两年在两台仪器上测量的38000个样本的基础上，我们现在表明这种方法非常稳健。在一个月内进行了多达1500次分析，并且可以在单个柱上分析> 14000个样品而不会损失色谱性能。样品包括细胞系、组织和人体体液的蛋白质组，其在有或没有先前肽分级分离或稳定同位素标记的情况下进行分析。我们表明，μ LC-MS/MS系统能够在1天内测量来自未耗尽的人血浆的2600种蛋白质和来自粗制人尿液的5000种蛋白质，表明其具有深入和高通量临床应用的潜力。
Microflow liquid chromatography tandem mass spectrometry (mu LC-MS/MS) is becoming a viable alternative to nanoflow LC-MS/MS for the analysis of proteomes. We have recently demonstrated the potential of such a system operating with a 1 mm i.d. X 150 mm column and at a flow rate of 50 mu L/ min for high-throughput applications. On the basis of the analysis of similar to 38 000 samples measured on two instruments over the past two years, we now show that the approach is extremely robust. Up to 1500 analyses were performed within one month, and >14 000 samples could be analyzed on a single column without loss of chromatographic performance. Samples included proteomes of cell lines, tissues, and human body fluids, which were analyzed with or without prior peptide fractionation or stable isotope labeling. We show that the mu LC-MS/MS system is capable of measuring 2600 proteins from undepleted human plasma and similar to 5000 proteins from crude human urine in 1 day, demonstrating its potential for in-depth as well as high-throughput clinical application.