MacCoss - Q-OT vs Q-LIT submission

Comparison of unit resolution versus high-resolution accurate mass for parallel reaction monitoring
Data License: CC BY 4.0 | ProteomeXchange: PXD023334 | doi: https://doi.org/10.6069/5b38-fg24
  • Organism: Homo sapiens
  • Instrument: Orbitrap Eclipse,Orbitrap Fusion Lumos
  • SpikeIn: No
  • Keywords: mass spectrometry, limit of quantitation, precision, PRM
  • Lab head: Michael MacCoss Submitter: Lilian Heil
Abstract
Parallel reaction monitoring (PRM) is an increasingly popular alternative to selected reaction monitoring (SRM) for targeted proteomics. PRM’s strengths over SRM are that it monitors all product ions in a single spectrum, thus eliminating the need to select interference-free product ions prior to data acquisition, and that it is most frequently performed on high-resolution instruments, such as quadrupole-orbitrap and quadrupole-time of flight instruments. Here, we show that the primary advantage of PRM is the ability to monitor all transitions in parallel, and that high-resolution data are not necessary to obtain high quality quantitative data. We run the same scheduled PRM assay, measuring 432 peptides from 126 plasma proteins, multiple times on a Orbitrap Eclipse Tribrid mass spectrometer, alternating separate liquid chromatography-tandem mass spectrometry runs between the high resolution Orbitrap and the unit resolution linear ion trap for PRM. We find that both mass analyzers have similar technical precision, and that the linear ion trap’s superior sensitivity gives it better lower limits of quantitation on over 62% of peptides in the assay.
Created on 4/28/21, 9:08 PM

All Skyline documents used for analysis are included in the Targeted MS Runs section and described briefly below:

Skyline document Description Raw data folder
qLITvsqOT_precision_final_2020-12-30_07-35-14.sky.zip Replicate injections in linear ion trap and Orbitrap using precursor isolation windows of 0.7 and 1.7 Th replicate_injections
qLITvsqOT_matrix_matched_originaltransitions_LIT_2021-04-27_14-24-59.sky.zip Matrix-matched dilution curves in linear ion trap using original transition set matrix_matched
qLITvsqOT_matrix_matched_originaltransitions_OT_2021-04-27_14-23-49.sky.zip Matrix-matched dilution curves in Orbitrap using original transition set matrix_matched
qLITvsqOT_matrix_matched_alltransitions_LIT_2021-04-27_14-13-12.sky.zip Matrix-matched dilution curves in linear ion trap with all possible transitions matrix_matched
qLITvsqOT_matrix_matched_alltransitions_OT_2021-04-27_14-18-08.sky.zip Matrix-matched dilution curves in Orbitrap with all possible transitions matrix_matched

Subsequent processing of the data was done externally, and with the code available here. Skyline documents include calibration curves, but limit of quantitation was determined externally with the method described by Pino et al. (2020).

Clustergrammer Heatmap
 
Download
20200704_ITvOT_plasma_final_2021-04-28_13-32-15.sky.zip2021-04-28 20:58:381544344342,984140
qLITvsqOT_matrix_matched_originaltransitions_LIT_2021-04-27_14-24-59.sky.zip2021-04-28 20:58:381244234232,95933423
qLITvsqOT_matrix_matched_originaltransitions_OT_2021-04-27_14-23-49.sky.zip2021-04-28 20:58:381244234232,95933423
qLITvsqOT_matrix_matched_alltransitions_OT_2021-04-27_14-18-08.sky.zip2021-04-28 20:58:381244234236,20233423
qLITvsqOT_matrix_matched_alltransitions_LIT_2021-04-27_14-13-12.sky.zip2021-04-28 20:58:381244234236,20233423
qLITvsqOT_precision_final_2020-12-30_07-35-14.sky.zip2021-04-28 20:58:381284604603,014200