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Mol Cell Proteomics. 2006 Jul;5(7):1338-47. Epub 2006 Mar 21.

Label-free quantitative proteomics using large peptide data sets generated by nanoflow liquid chromatography and mass spectrometry.

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1
Chemotherapy Division and Cancer Proteomics Project, National Cancer Center Research Institute, Tokyo 104-0045, Japan.

Abstract

We developed an integrated platform consisting of machinery and software modules that can apply vast amounts of data generated by nanoflow LC-MS to differential protein expression analyses. Unlabeled protein samples were completely digested with modified trypsin and separated by low speed (200 nl/min) one-dimensional HPLC. Mass spectra were obtained every 1 s by using the survey mode of a hybrid Q-TOF mass spectrometer and displayed in a two-dimensional plane with m/z values along the x axis, and retention time was displayed along the y axis. The time jitter of nano-LC was adjusted using newly developed software based on a dynamic programming algorithm. The comprehensiveness (60,000-160,000 peaks above the predetermined threshold detectable in 60-microg cell protein samples), reproducibility (average coefficient of variance of 0.35-0.39 and correlation coefficient of over 0.92 between duplicates), and accurate quantification with a wide dynamic range (over 10(3)) of our platform warrant its application to various types of experimental and translational proteomics.

PMID:
16552026
DOI:
10.1074/mcp.T500039-MCP200
[Indexed for MEDLINE]
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