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J Proteome Res. 2016 Mar 4;15(3):691-706. doi: 10.1021/acs.jproteome.5b00859. Epub 2015 Dec 22.

Reproducibility of Differential Proteomic Technologies in CPTAC Fractionated Xenografts.

Author information

1
Department of Mathematical Sciences, University of Cincinnati , Cincinnati, Ohio 45221, United States.
2
Proteomics Platform, Broad Institute of MIT and Harvard , Cambridge, Massachusetts 02142, United States.
3
Department of Biochemistry and Biophysics, University of North Carolina , Chapel Hill, North Carolina 27599, United States.
4
Department of Medicine, Washington University , St. Louis, Missouri 63110, United States.
5
Office of Cancer Clinical Proteomics Research, National Cancer Institute , Bethesda, Maryland 20892, United States.
6
Enterprise Science and Computing, Inc. , Rockville, Maryland 20850, United States.
7
Division of Biological Sciences, Pacific Northwest National Laboratory , Richland, Washington 99352, United States.
8
JHMI and Division of Clinical Chemistry, Johns Hopkins University , Baltimore, Maryland 21231, United States.

Abstract

The NCI Clinical Proteomic Tumor Analysis Consortium (CPTAC) employed a pair of reference xenograft proteomes for initial platform validation and ongoing quality control of its data collection for The Cancer Genome Atlas (TCGA) tumors. These two xenografts, representing basal and luminal-B human breast cancer, were fractionated and analyzed on six mass spectrometers in a total of 46 replicates divided between iTRAQ and label-free technologies, spanning a total of 1095 LC-MS/MS experiments. These data represent a unique opportunity to evaluate the stability of proteomic differentiation by mass spectrometry over many months of time for individual instruments or across instruments running dissimilar workflows. We evaluated iTRAQ reporter ions, label-free spectral counts, and label-free extracted ion chromatograms as strategies for data interpretation (source code is available from http://homepages.uc.edu/~wang2x7/Research.htm ). From these assessments, we found that differential genes from a single replicate were confirmed by other replicates on the same instrument from 61 to 93% of the time. When comparing across different instruments and quantitative technologies, using multiple replicates, differential genes were reproduced by other data sets from 67 to 99% of the time. Projecting gene differences to biological pathways and networks increased the degree of similarity. These overlaps send an encouraging message about the maturity of technologies for proteomic differentiation.

KEYWORDS:

CPTAC; Differential proteomics; iTRAQ; label-free; quality control; technology assessment; xenografts

PMID:
26653538
PMCID:
PMC4779376
DOI:
10.1021/acs.jproteome.5b00859
[Indexed for MEDLINE]
Free PMC Article

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