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High‐sensitivity analysis and sequencing of peptides and proteins by quadrupole ion trap mass spectrometry
Author(s) -
Marina Anabel,
García Miguel Angel,
Albar Juan Pablo,
Yagüe Jesús,
López de Castro José Antonio,
Vázquez Jesús
Publication year - 1999
Publication title -
journal of mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 121
eISSN - 1096-9888
pISSN - 1076-5174
DOI - 10.1002/(sici)1096-9888(199901)34:1<17::aid-jms746>3.0.co;2-6
Subject(s) - chemistry , ion trap , mass spectrometry , quadrupole ion trap , electrospray , chromatography , analytical chemistry (journal) , deconvolution , fragmentation (computing) , ion , mass spectrum , algorithm , computer science , organic chemistry , operating system
This paper describes experience with the commercially available LCQquadrupole ion trap mass spectrometer applied to the off‐lineanalysis of peptides and proteins. The standard front end of theelectrospray probe was replaced with a micromanipulator which, withthe aid of a magnifying device, allowed the use of a variety ofminiaturized spraying interfaces. The low sample consumption andextended analysis times of these devices were ideally suitable toobtain improved results in terms of sensitivity and mass accuracy.This needed a careful optimization of the number of ions storedinside the trap (ion target parameter) and requiredspectrum averaging of many scans. A method is presented for themathematical fitting of ZoomScan spectra to theoretical isotopicdistributions, which allowed the mass determination of large peptideswith more accuracy than that achieved by conventional deconvolutionalgorithms. A very simple on‐line desalting configuration isalso described which needed no externalmicro‐high‐performance liquid chromatographic pumps,and can be easily mounted using the built‐in syringe deliverysystem of the LCQ. This set‐up allowed extended analysis timesof ‘in‐gel’ protein digests in subpicomoleamounts. Finally, the multiple fragmentation capabilities of the iontrap were found to be extremely useful for the analysis of peptidemodifications such as phosphorylation and for sequencing individualpeptides from highly complex MHC‐bound peptide pools.Copyright © 1999 John Wiley & Sons, Ltd.