A Compact Quadrupole-Orbitrap Mass Spectrometer with FAIMS Interface Improves Proteome Coverage in Short LC Gradients
A Compact Quadrupole-Orbitrap Mass Spectrometer with FAIMS Interface Improves Proteome Coverage in Short LC Gradients
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DOI：
10.1074/mcp.tir119.001906
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发表时间：
2020-04-01
影响因子：
7
通讯作者：
Olsen, Jesper, V
中科院分区：
文献类型：
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作者：
Bekker-Jensen, Dorte B.;Martinez-Val, Ana;Olsen, Jesper, V
最先进的蛋白质组学级质谱仪可以以每秒数十个肽的测序速度和阿摩尔灵敏度以ppm质量精度测量肽前体及其片段。在这里，我们描述了一个紧凑和强大的四极轨道阱质谱仪配备了前端高场非对称波形离子迁移谱（FAIMS）接口。结合FAIMS，在数据相关采集（DDA）和数据无关采集（DIA）模式下评估了Orbitrap Exploris 480质谱仪的性能。我们证明，不同的补偿电压（CV）FAIMS是最佳的DDA和DIA，分别。使用单个CV将DIA与FAIMS相结合，该仪器每分钟识别超过2500个肽。这使得通过Evosep One LC系统提供的短在线LC梯度能够定量>5000种蛋白质，允许每天采集60个样品。通过分析5 ng HeLa消化物来评估仪器的原始灵敏度，其中使用DIA-FAIMS用5分钟LC梯度可重复地鉴定出>1000种蛋白质。为了证明该仪器的多功能性，我们记录了12个大鼠组织中蛋白质组表达的器官范围图，该蛋白质组表达通过串联质量标签和使用DIA与FAIMS的无标记定量进行定量，深度> 10，000个蛋白质。
State-of-the-art proteomics-grade mass spectrometers can measure peptide precursors and their fragments with ppm mass accuracy at sequencing speeds of tens of peptides per second with attomolar sensitivity. Here we describe a compact and robust quadrupole-orbitrap mass spectrometer equipped with a front-end High Field Asymmetric Waveform Ion Mobility Spectrometry (FAIMS) Interface. The performance of the Orbitrap Exploris 480 mass spectrometer is evaluated in data-dependent acquisition (DDA) and data-independent acquisition (DIA) modes in combination with FAIMS. We demonstrate that different compensation voltages (CVs) for FAIMS are optimal for DDA and DIA, respectively. Combining DIA with FAIMS using single CVs, the instrument surpasses 2500 peptides identified per minute. This enables quantification of >5000 proteins with short online LC gradients delivered by the Evosep One LC system allowing acquisition of 60 samples per day. The raw sensitivity of the instrument is evaluated by analyzing 5 ng of a HeLa digest from which >1000 proteins were reproducibly identified with 5 min LC gradients using DIA-FAIMS. To demonstrate the versatility of the instrument, we recorded an organ-wide map of proteome expression across 12 rat tissues quantified by tandem mass tags and label-free quantification using DIA with FAIMS to a depth of >10,000 proteins.