title: A comparison of nLC-ESI-MS/MS and nLC-MALDI-MS/MS for GeLC-based protein identification and iTRAQ-based shotgun quantitative proteomics.
abstract: The use of nLC-ESI-MS/MS in shotgun proteomics experiments and GeLC-MS/MS analysis is well accepted and routinely available in most proteomics laboratories. However, the same cannot be said for nLC-MALDI MS/MS, which has yet to experience such widespread acceptance, despite the fact that the MALDI technology offers several critical advantages over ESI. As an illustration, in an analysis of moderately complex sample of E. coli proteins, the use MALDI in addition to ESI in GeLC-MS/MS resulted in a 16% average increase in protein identifications, while with more complex samples the number of additional protein identifications increased by an average of 45%. The size of the unique peptides identified by MALDI was, on average, 25% larger than the unique peptides identified by ESI, and they were found to be slightly more hydrophilic. The insensitivity of MALDI to the presence of ionization suppression agents was shown to be a significant advantage, suggesting it be used as a complement to ESI when ion suppression is a possibility. Furthermore, the higher resolution of the TOF/TOF instrument improved the sensitivity, accuracy, and precision of the data over that obtained using only ESI-based iTRAQ experiments using a linear ion trap. Nevertheless, accurate data can be generated with either instrument. These results demonstrate that coupling nanoLC with both ESI and MALDI ionization interfaces improves proteome coverage, reduces the deleterious effects of ionization suppression agents, and improves quantitation, particularly in complex samples.
authors.affiliation: U.S. Plant, Soil & Nutrition Laboratory, Tower Road, Ithaca, NY 14853, USA.
authors.forename: Yong
authors.initials: Y
authors.lastname: Yang
authors.forename: Sheng
authors.initials: S
authors.lastname: Zhang
authors.forename: Kevin
authors.initials: K
authors.lastname: Howe
authors.forename: David B
authors.initials: DB
authors.lastname: Wilson
authors.forename: Felix
authors.initials: F
authors.lastname: Moser
authors.forename: Diana
authors.initials: D
authors.lastname: Irwin
authors.forename: Theodore W
authors.initials: TW
authors.lastname: Thannhauser
mesh_terms: D000595:Amino Acid Sequence; D002853:Chromatography, Liquid / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D036103:Nanotechnology; D040901:Proteomics