用于相对和绝对定量的同量异位标签（iTRAQ）中的技术、实验和生物学变异。

Technical, experimental, and biological variations in isobaric tags for relative and absolute quantitation (iTRAQ).

作者信息

Gan Chee Sian, Chong Poh Kuan, Pham Trong Khoa, Wright Phillip C

机构信息

Biological and Environmental Systems Group, Department of Chemical and Process Engineering, The University of Sheffield, Mappin Street, Sheffield S1 3JD, United Kingdom.

出版信息

J Proteome Res. 2007 Feb;6(2):821-7. doi: 10.1021/pr060474i.

DOI:10.1021/pr060474i

PMID:17269738

Abstract

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.

摘要

我们基于不同类型的重复分析，考虑技术、实验和生物学变异，评估用于相对和绝对定量的等压标签（iTRAQ）的可靠性。总共对涉及酿酒酵母KAY446、嗜热栖热菌P2和聚球藻属PCC 6803的三个生命领域的10个iTRAQ实验进行了分析。随着变异容忍度的增加，iTRAQ分析的蛋白质表达覆盖率也会增加。简而言之，基于生物学重复分析，在变异为±50%（±0.50）的截止点下，定量覆盖率将达到88%。技术重复分析在较低的±30%变异截止点下可产生95%的更高覆盖率。实验或iTRAQ变异与生物学变异表现出相似的行为，这表明大多数可测量的偏差来自生物学变异。这些发现强调了重复分析作为蛋白质表达分析中的验证工具和基准技术的重要性。