Vincent Coates Foundation Mass Spectrometry Laboratory, Stanford University, Stanford, CA, 94305, USA.
Sean Parker Center, Stanford University School of Medicine, Stanford, CA, 94305, USA.
Proteomics. 2020 Jun;20(11):e1900105. doi: 10.1002/pmic.201900105. Epub 2020 Mar 3.
The analytical scale of most mass-spectrometry-based targeted proteomics assays is usually limited by assay performance and instrument utilization. A recently introduced method, called triggered by offset, multiplexed, accurate mass, high resolution, and absolute quantitation (TOMAHAQ), combines both peptide and sample multiplexing to simultaneously improve analytical scale and quantitative performance. In the present work, critical technical requirements and data analysis considerations for successful implementation of the TOMAHAQ technique based on the study of a total of 185 target peptides across over 200 clinical plasma samples are discussed. Importantly, it is observed that significant interference originate from the TMTzero reporter ion used for the synthetic trigger peptides. This interference is not expected because only TMT10plex reporter ions from the target peptides should be observed under typical TOMAHAQ conditions. In order to unlock the great promise of the technique for high throughput quantification, here a post-acquisition data correction strategy to deconvolute the reporter ion superposition and recover reliable data is proposed.
基于质谱的靶向蛋白质组学分析的分析规模通常受到分析方法性能和仪器利用率的限制。最近引入的一种方法称为偏移触发、多重化、精确质量、高分辨率和绝对定量（TOMAHAQ），它结合了肽和样品的多重化，以同时提高分析规模和定量性能。在本工作中，基于对超过 200 个临床血浆样本中的 185 个靶肽的研究，讨论了成功实施 TOMAHAQ 技术的关键技术要求和数据分析注意事项。重要的是，观察到来自用于合成触发肽的 TMTzero 报告离子的显著干扰。这种干扰是不期望的，因为在典型的 TOMAHAQ 条件下，仅应观察到来自目标肽的 TMT10plex 报告离子。为了释放该技术在高通量定量方面的巨大潜力，本文提出了一种在采集后进行数据校正的策略，以解卷积报告离子叠加并恢复可靠的数据。
