Technical, experimental, and biological variations in isobaric tags for relative and absolute quantitation (iTRAQ)
Technical, experimental, and biological variations in isobaric tags for relative and absolute quantitation (iTRAQ)
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DOI：
10.1021/pr060474i
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发表时间：
2007-02-01
影响因子：
4.4
通讯作者：
Wright, Phillip C.
中科院分区：
文献类型：
--
作者：
Gan, Chee Sian;Chong, Poh Kuan;Wright, Phillip C.
我们评估相对和绝对定量等压标签（iTRAQ）的可靠性，基于不同类型的重复分析，考虑到技术、实验和生物变异。共对10个iTRAQ实验进行了3个生命域的分析，涉及酿酒酵母KAY446、solfataricus Sulfolobus P2和Synechocystis sp. PCC 6803。随着变异耐受性的增加，iTRAQ分析的蛋白表达覆盖率增加。简而言之，在基于生物重复分析的定量分析中，+/- 50%变异（+/- 0.50）的截断点将产生88%的覆盖率。技术重复分析在+/- 30%变化的较低截止点上产生了95%的较高覆盖率。实验或iTRAQ变异表现出与生物变异相似的行为，这表明大多数可测量的偏差来自生物变异。这些发现强调了重复分析作为蛋白质表达分析的验证工具和基准技术的重要性。
We assess the reliability of isobaric-tags for relative and absolute quantitation (iTRAQ), based on different types of replicate analyses taking into account technical, experimental, and biological variations. In total, 10 iTRAQ experiments were analyzed across three domains of life involving Saccharomyces cerevisiae KAY446, Sulfolobus solfataricus P2, and Synechocystis sp. PCC 6803. The coverage of protein expression of iTRAQ analysis increases as the variation tolerance increases. In brief, a cutoff point at +/- 50% variation (+/- 0.50) would yield 88% coverage in quantification based on an analysis of biological replicates. Technical replicate analysis produces a higher coverage level of 95% at a lower cutoff point of +/- 30% variation. Experimental or iTRAQ variations exhibit similar behavior as biological variations, which suggest that most of the measurable deviations come from biological variations. These findings underline the importance of replicate analysis as a validation tool and benchmarking technique in protein expression analysis.