Using Data-Independent Mass Spectrometry to Extend Detectable Dynamic Range without Prior Fractionation - Proteomic and Metabolomic Approaches to Biomarker Discovery - page 120

FIGURE 2 Quantitative PAcIFIC acquisition using tandem mass tag (TMT). (A) In between each regular CID scan
used for identi cation, a narrow PQD scan is acquired only in the m/z range corresponding to the reporter ions for

faster scanning time. (B) Both CID and PQD scans are merged to create a composite spectrum that is further used for
database searching and quanti cation. (Reprinted with permission from reference #16. Copyright 2011 American Chemical

Society.)

P. aeruginosa, it was still effective at identifying
and quantifying a large number of proteins. 16

In addition, these orphan peptides are detectable
fromMS2 but not MS1 scans in our DIA PAcIFIC
experiments because of the intrinsically higher
signal to noise for tandem mass spectra than
precursor ion mass spectra. We also investigated
whether there was a difference in occurrence
of accidental CID events (i.e., co-fragmenting
peptides, a.k.a. chimeric peptides) between the
PAcIFIC DIA and standard DDA protocols. 11,16
In order to determine the extent to which
these two categories occurred in an optimized
PAcIFIC strategy, we developed a modi

PAcIFIC with High-Resolution High
Mass Accuracy Precursor Ion Scans
Interestingly, using the DIA PAcIFIC anal-
ysis, but not DDA methods, allowed us to detect
a class of peptides for which no precursor ions
could be detected, which we refer to as

“ orphan ”

peptides. These orphan peptides were identi ed

solely based on the quality of their tandem mass
spectra and were a result of selecting ions from
m/z channels in the absence of precursor ions.

ed
PAcIFIC method in which high mass accuracy
precursor ion mass spectra were also acquired.