Journal
ANALYTICAL BIOCHEMISTRY
Volume 560, Issue -, Pages 39-49Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ab.2018.08.025
Keywords
Tryptic peptides; Random and independent sampling; LC-ESI-MS/MS; Chi square; ANOVA; Type I error rate
Funding
- Natural Science and Engineering Research Council of Canada (NSERC)
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A Rabbit myosin standard, like that used to create the empirical statistical model, was randomly and independently sampled by liquid chromatography micro electrospray ionization and tandem mass spectrometry (LC-ESI-MS/MS) with a linear quadrupole ion trap. The rabbit myosin protein standard appeared pure by SDS-PAGE and CBBR staining but showed many other proteins by silver staining. The LC-MS intensity from myosin and IgG samples were above the 99% safe limit of detection and quantification computed from 36 blank LC-ESI-MS/MS runs. The myosin contained <= 406 Gene Symbols, open reading frames or loci where 79 protein types showed >= 3 peptides from X!TANDEM. Myosins, actin, troponin, other proteins showed 95%-100% homology between the rabbit versus the human decoy library. The myosin protein complex from STRING was true positive compared to random or noise spectra MS/MS with a low type I error (p-value) and low FDR (q-value) computed in R. SDS-PAGE, Western blot, comparison to random and noise MS/MS spectra, X!TANDEM p-values, FDR corrected q-values, and STRING all agreed that the error rate of LC-ESI-MS/MS with a quadrupole ion trap is far below that assumed a priori by the design of the empirical statistical model for decoy library searching.
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