Peptide Labeling with Isobaric Tags Yields Higher Identification Rates Using iTRAQ 4-Plex Compared to TMT 6-Plex and iTRAQ 8-Plex on LTQ Orbitrap
In: Analytical chemistry (Washington), Jg. 82 (2010), Heft 15, S. 6549-6558
academicJournal
- print,
Zugriff:
Peptide labeling with isobaric tags has become a popular technique in quantitative shotgun proteomics. Using two different samples viz. a protein mixture and HeLa extracts, we show that three commercially available isobaric tags differ with regard to peptide identification rates: The number of identified proteins and peptides was largest with iTRAQ 4-plex, followed by TMT 6-plex, and smallest with iTRAQ 8-plex. In all experiments, we employed a previously described method where two scans were acquired for each precursor on an LTQ Orbitrap: A CID scan under standard settings for identification, and a HCD scan for quantification. The observed differences in identification rates were similar when data was searched with either Mascot or Sequest. We consider these findings to be the result of a combination of several factors, most notably prominent ions in CID spectra as a consequence of loss of fragments of the label tag from precursor ions. These fragment ions cannot be explained by current search engines and were observed to have a negative impact on peptide scores.
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Peptide Labeling with Isobaric Tags Yields Higher Identification Rates Using iTRAQ 4-Plex Compared to TMT 6-Plex and iTRAQ 8-Plex on LTQ Orbitrap
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Autor/in / Beteiligte Person: | PICHLER, Peter ; KÖCHER, Thomas ; HOLZMANN, Johann ; MAZANEK, Michael ; TAUS, Thomas ; AMMERER, Gustav ; MECHTLER, Karl |
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Zeitschrift: | Analytical chemistry (Washington), Jg. 82 (2010), Heft 15, S. 6549-6558 |
Veröffentlichung: | Washington, DC: American Chemical Society, 2010 |
Medientyp: | academicJournal |
Umfang: | print, |
ISSN: | 0003-2700 (print) |
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