Journal
JOURNAL OF PROTEOME RESEARCH
Volume 12, Issue 2, Pages 594-604Publisher
AMER CHEMICAL SOC
DOI: 10.1021/pr300624g
Keywords
Mass spectrometry; iTRAQ; statistical analysis; experimental design
Categories
Funding
- Bill and Melinda Gates Foundation [GH 5241, GH614]
- US Agency for International Development, Washington, D.C. [HRN-A-00-97-00015-00, GHS-A-00-03-00019-00]
- Johns Hopkins University
- Bill and Melinda Gates Foundation, Seattle, WA [GH 614]
- CTSA grant
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Isobaric tags for relative and absolute quantitation (iTRAQ) is a prominent mass spectrometry technology for protein identification and quantification that is capable of analyzing multiple samples in a single experiment. Frequently, iTRAQ experiments are carried out using an aliquot from a pool of all samples, or masterpool, in one of the channels as a reference sample standard to estimate protein relative abundances in the biological samples and to combine abundance estimates from multiple experiments. In this manuscript, we show that using a masterpool is counterproductive. We obtain more precise estimates of protein relative abundance by using the available biological data instead of the masterpool and do not need to occupy a channel that could otherwise be used for another biological sample. In addition, we introduce a simple statistical method to associate proteomic data from multiple iTRAQ experiments with a numeric response and show that this approach is more powerful than the conventionally employed masterpool-based approach. We illustrate our methods using data from four replicate iTRAQ experiments on aliquots of the same pool of plasma samples and from a 406-sample project designed to identify plasma proteins that covary with nutrient concentrations in chronically undernourished children from South Asia.
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