Simultaneous Proteomic Discovery and Targeted Monitoring using Liquid Chromatography, Ion Mobility Spectrometry, and Mass Spectrometry
Simultaneous Proteomic Discovery and Targeted Monitoring using Liquid Chromatography, Ion Mobility Spectrometry, and Mass Spectrometry
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DOI：
10.1074/mcp.m116.061143
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发表时间：
2016-12-01
影响因子：
7
通讯作者：
Smith，Richard D.
中科院分区：
文献类型：
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作者：
Burnum-Johnson，Kristin E.;Nie，Song;Smith，Richard D.
当前的蛋白质组学方法包括广泛的发现测量和定量目标分析。在许多情况下，发现测量最初用于识别潜在重要的蛋白质（例如候选生物标志物），然后采用有针对性的研究来量化有限数量的选定蛋白质。然而，这两种方法都存在局限性。发现测量旨在对整个蛋白质组进行采样，但其灵敏度、准确性和定量精度低于靶向方法，而靶向测量的灵敏度明显更高，但仅对蛋白质组的有限部分进行采样。在此，我们描述了一种新方法，通过结合液相色谱、离子迁移谱和质谱 (LC-IMS-MS) 在一次分析中执行发现和目标监测 (DTM)。在 DTM 中，重标记的目标肽被掺入胰蛋白酶消化物中，并使用 LC-IMS-MS 仪器检测标记和未标记的肽。与广泛的 LC-MS 发现测量相比，DTM 产生更大的肽/蛋白质覆盖率并检测较低丰度的物种。 DTM 还达到了与选择反应监测 (SRM) 类似的检测限，表明其将高质量发现和目标分析相结合的潜力，这是朝着发现和目标方法融合迈出的重要一步。
Current proteomic approaches include both broad discovery measurements and quantitative targeted analyses. In many cases, discovery measurements are initially used to identify potentially important proteins (e.g.candidate biomarkers) and then targeted studies are employed to quantify a limited number of selected proteins. Both approaches, however, suffer from limitations. Discovery measurements aim to sample the whole proteome but have lower sensitivity, accuracy, and quantitation precision than targeted approaches, whereas targeted measurements are significantly more sensitive but only sample a limited portion of the proteome. Herein, we describe a new approach that performs both discovery and targeted monitoring (DTM) in a single analysis by combining liquid chromatography, ion mobility spectrometry and mass spectrometry (LC-IMS-MS). In DTM, heavy labeled target peptides are spiked into tryptic digests and both the labeled and unlabeled peptides are detected using LC-IMS-MS instrumentation. Compared with the broad LC-MS discovery measurements, DTM yields greater peptide/protein coverage and detects lower abundance species. DTM also achieved detection limits similar to selected reaction monitoring (SRM) indicating its potential for combined high quality discovery and targeted analyses, which is a significant step toward the convergence of discovery and targeted approaches.