Mt. Davis et al., Towards defining the urinary proteome using liquid chromatography-tandem mass spectrometry - II. Limitations of complex mixture analyses, PROTEOMICS, 1(1), 2001, pp. 108-117
With an emphasis on obtaining a multitude of high quality tandem mass spect
rometry spectra for protein identification, instrumental parameters are des
cribed for the liquid chromatography-tandem mass spectrometry analysis of t
rypsin digested unfractionated urine using a hybrid quadrupole-time-of-flig
ht (Q-TOF) mass spectrometer. Precursor acquisition rates of up to 20 disti
nct precursors/minute in a single analysis were obtained through the use of
parallel precursor selection (four precursors/survey period) and variable
collision induced dissociation integration time (1 to 6 periods summed). Ma
ximal exploitation of the gas phase fractionated ions was obtained through
the use of narrow survey scans and iterative data-dependent analyses incorp
orating dynamic exclusion. The impact on data fidelity as a product of data
-dependent selection of precursor ions from a dynamically excluded field is
discussed with regards to sample complexity, precursor selection rates, su
rvey scan range and facile chemical modifications. Operational and post-ana
lysis strategies are presented to restore data confidence and reconcile the
greatest number of matched spectra.