A Strategy to Combine Sample Multiplexing with Targeted Proteomics Assays for High-Throughput Protein Signature Characterization.
A Strategy to Combine Sample Multiplexing with Targeted Proteomics Assays for High-Throughput Protein Signature Characterization.
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一种将样品多路复用与靶向蛋白质组学测定法相结合的策略，以进行高通量蛋白质特征。
DOI：
10.1016/j.molcel.2016.12.005
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发表时间：
2017-01-19
期刊：
影响因子：
16
通讯作者：
Gygi SP
中科院分区：
文献类型：
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作者：
Erickson BK;Rose CM;Braun CR;Erickson AR;Knott J;McAlister GC;Wühr M;Paulo JA;Everley RA;Gygi SP
用于蛋白质定量的靶向质谱分析监测肽替代物，其易于在单个分析中多重靶向许多肽。然而，这些测定通常没有利用允许平行进行多达10次分析的样品多路复用。我们提出了一种二维多路复用工作流程，该工作流程利用每种蛋白质的合成肽来促进来自多达10种混合样品条件的>100种肽的同时定量。我们证明，有针对性的分析未分级裂解物（2小时）准确再现分级裂解物（72小时分析）的定量，同时避免了肽检测之前的定量需要。我们在所有60个国家癌症研究所细胞系中靶向了对应于69种蛋白质的131种肽，一式三份，仅在48小时内分析了180个样品（相当于16分钟/样品）。这些数据进一步阐明了关键蛋白质的表达与其对药物治疗的细胞反应之间的相关性。
Targeted mass spectrometry assays for protein quantitation monitor peptide surrogates, which are easily multiplexed to target many peptides in a single assay. However, these assays have generally not taken advantage of sample multiplexing which allows up to 10 analyses to occur in parallel. We present a two-dimensional multiplexing workflow that utilizes synthetic peptides for each protein to prompt the simultaneous quantification of >100 peptides from up to 10 mixed sample conditions. We demonstrate that targeted analysis of unfractionated lysates (2-hr) accurately reproduces the quantification of fractionated lysates (72-hr analysis), while obviating the need for peptide detection prior to quantification. We targeted 131 peptides corresponding to 69 proteins across all 60 National Cancer Institute cell lines in biological triplicate, analyzing 180 samples in only 48 hours (the equivalent of 16 min/sample). These data further elucidated a correlation between the expression of key proteins and their cellular response to drug treatment.