Liquid chromatography–mass spectrometry (LC–MS) delivers sensitive peptide analysis for proteomics but requires extensive analysis time, reducing throughput. Here, we demonstrate that gas-phase peptide separation instead of LC enables fast proteome analysis. Using direct infusion–shotgun proteome analysis (DI-SPA) by data-independent acquisition mass spectrometry (DIA-MS), we demonstrate the targeted quantification of over 500 proteins within minutes of MS data collection (~3.5 proteins per second). We show the utility of this technology in performing a complex multifactorial proteomic study of interactions between nutrients, genotype and mitochondrial toxins in a collection of cultured human cells. More than 45,000 quantitative protein measurements from 132 samples were achieved in only ~4.4 h of MS data collection. Enabling fast, unbiased proteome quantification without LC, DI-SPA offers an approach to boost throughput, critical to drug and biomarker discovery studies that require analysis of thousands of proteomes.

液相色谱-质谱（LC-MS）可以对蛋白质组学进行灵敏的肽段分析，但需要大量的分析时间，从而降低了通量。在这里，我们证明了气相肽分离而不是液相色谱可以实现快速蛋白质组分析。使用直接输注-shot弹枪蛋白质组分析（DI-SPA）和独立于数据的采集质谱仪（DIA-MS），我们证明了在MS数据收集的几分钟内可以对500多种蛋白质进行靶向定量（每秒约3.5个蛋白质）。我们展示了这项技术在执行人类细胞集合中营养，基因型和线粒体毒素之间相互作用的复杂多因素蛋白质组学研究中的实用性。仅在约4.4小时的MS数据收集中，就从132个样品中进行了超过45,000个定量蛋白质测量。快速启用