Document Detail


A study of reproducibility of guanidination-dimethylation labeling and liquid chromatography matrix-assisted laser desorption ionization mass spectrometry for relative proteome quantification.
MedLine Citation:
PMID:  17386668     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The combination of dimethylation after guanidination (2MEGA) isotope labeling with microbore liquid chromatography (LC)-matrix-assisted laser desorption ionization (MALDI) MS and MS/MS [C. Ji, N. Guo, L. Li, J. Proteome Res. 4 (2005) 2099] has been reported as a promising strategy for abundance ratio-dependent quantitative proteome analysis. A critical step in using this integrated strategy is to set up the abundance ratio threshold of peptide pairs, above which the peptide pairs are used for quantifying and identifying the protein that is considered to be differentially expressed between two different samples. The threshold is determined by technical variation (i.e., the overall abundance ratio variation caused by the experimental process including sample workup, MS analysis and data processing) as well as biological variation (i.e., the abundance ratio variation caused by the biological process including cell growth), which can be defined and assessed by a coefficient of variation (CV). We have designed experiments and measured three different levels of variations, starting with the same membrane protein preparation, the same batch of cells and three batches of cells from the same cell line grown under the same conditions, respectively. It is shown that technical variation from the experimental processes involved in 2MEGA labeling LC-MALDI MS has a CV of <15%. In addition, the measured biological variation from cell growth was much smaller than the measured technical variation. From the studies of the occurrence rate of outliers in the distribution of the abundance ratio data within a comparative dataset of peptide pairs, it is concluded that, to compare the proteome changes between two sets of cultured cells without the use of replicate experiments, a relative abundance ratio of greater than 2X or less than 0.5X (X is the average abundance ratio of the dataset) on peptide pairs can be used as a stringent threshold to quantify and identify differentially expressed proteins with high confidence.
Authors:
Chengjie Ji; Nan Zhang; Sambasivarao Damaraju; Vijaya L Damaraju; Pat Carpenter; Carol E Cass; Liang Li
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2007-01-13
Journal Detail:
Title:  Analytica chimica acta     Volume:  585     ISSN:  1873-4324     ISO Abbreviation:  Anal. Chim. Acta     Publication Date:  2007 Mar 
Date Detail:
Created Date:  2007-03-27     Completed Date:  2007-07-06     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  0370534     Medline TA:  Anal Chim Acta     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  219-26     Citation Subset:  IM    
Affiliation:
Department of Chemistry, University of Alberta, Cross Cancer Institute, Edmonton, Alberta, Canada T6G 2G2.
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MeSH Terms
Descriptor/Qualifier:
Animals
Automation
Cell Line
Cell Line, Tumor
Cell Proliferation
Chromatography, Ion Exchange
Chromatography, Liquid / methods*
Guanidine / chemistry*
Humans
Methylation
Peptides / chemistry
Proteome
Proteomics / methods*
Reproducibility of Results
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / methods*
Chemical
Reg. No./Substance:
0/Peptides; 0/Proteome; 113-00-8/Guanidine

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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