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Towards defining the urinary proteome using liquid chromatography‐tandem mass spectrometry I.Profiling an unfractionated tryptic digest
Author(s) -
Spahr Chris S.,
Davis Michael T.,
McGinley Michael D.,
Robinson John H.,
Bures Edward J.,
Beierle Jill,
Mort Jessica,
Courchesne Paul L.,
Chen Kui,
Wahl Robert C.,
Yu Wen,
Luethy Roland,
Patterson Scott D.
Publication year - 2001
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/1615-9861(200101)1:1<93::aid-prot93>3.0.co;2-3
Subject(s) - chromatography , proteome , tandem mass spectrometry , mass spectrometry , liquid chromatography–mass spectrometry , chemistry , urinary system , proteomics , medicine , biochemistry , gene
The proteome of normal male urine from a commercial pooled source has been examined using direct liquid chromatography‐tandem mass spectrometry (LC‐MS/MS). The entire urinary protein mixture was denatured, reduced and enzymatically digested prior to LC‐MS/MS analysis using a hybrid‐quadrupole time‐of‐flight mass spectrometer (Q‐TOF) to perform data‐dependent ion selection and fragmentation. To fragment as many peptides as possible, the mixture was analyzed four separate times, with the mass spectrometer selecting ions for fragmentation from a subset of the entire mass range for each run. This approach requires only an autosampler on the HPLC for automation ( i.e , unattended operation). Across these four analyses, 1.450 peptide MS/MS spectra were matched to 751 sequences to identify 124 gene products (proteins and translations of expressed sequence tags). Interestingly, the experimental time for these analyses was less than that required to run a single two‐dimensional gel.