Repository logo
 

Prior Signal Acquisition Software Versions for Orbitrap Underestimate Low Isobaric Mass Tag Intensities, Without Detriment to Differential Abundance Experiments

Accepted version
Peer-reviewed

No Thumbnail Available

Type

Article

Change log

Authors

Andrejeva, Anna 
Crook, Oliver 
Elzek, Mohammed 

Abstract

Tandem mass tags (TMT) enable simple and accurate quantitative proteomics for multiplexed samples by relative quantification of tag reporter ions. Orbitrap™ quantification of reporter ions has been associated with a characteristic notch region in intensity distribution, within which few reporter intensities are recorded. This has been resolved in version 3 of the instrument acquisition software, Tune. However, 53 % of Orbitrap Fusion, Lumos or Eclipse submissions to PRIDE were generated using prior software versions. To quantify the impact of the notch on existing quantitative proteomics data, we generated a mixed species benchmark and acquired quantitative data using Tune versions 2 and 3. Sub-notch intensities are predominantly underestimated with Tune version 2, leading to overestimation of the true differences in intensities between samples. However, when summarising reporter ion intensities to higher level features, such as peptides and proteins, few features are significantly affected. Targeted removal of spectra with reporter ion intensities below the notch is not beneficial for differential peptide or protein testing. Overall, we find the systematic quantification bias associated with the notch is not detrimental for a typical proteomics experiment.

Description

Keywords

Journal Title

ACS Measurement Science Au

Conference Name

Journal ISSN

2694-250X
2694-250X

Volume Title

Publisher

American Chemical Society
Sponsorship
Wellcome Trust (110170/Z/15/Z)
Biotechnology and Biological Sciences Research Council (BB/R505304/1)
Wellcome Trust (110071/Z/15/Z)
European Commission Horizon 2020 (H2020) Research Infrastructures (RI) (823839)
Relationships
Is supplemented by: