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Related Experiment Video

Updated: Nov 12, 2025

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Strain-Specific Peptide (SSP) Interference Reference Sample: A Genetically Encoded Quality Control for Isobaric

Tian Zhang1, Greg R Keele2, Gary A Churchill2

  • 1Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, United States.

Analytical Chemistry
|March 18, 2021
PubMed
Summary

A new reference sample using mouse strain genetic variation helps quantify interference in tandem mass tag (TMT) proteomics. This tool improves the accuracy of protein abundance profiling by assessing the interference-free index (IFI).

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Area of Science:

  • Proteomics
  • Analytical Chemistry
  • Biochemistry

Background:

  • Isobaric tag-based multiplexing is crucial for global protein quantification.
  • Co-eluting peptide interference confounds these analyses, leading to ratio compression.
  • Existing methods like ion mobility offer partial solutions but require further validation.

Purpose of the Study:

  • To introduce a novel quality control sample for assessing interference in tandem mass tag (TMT) experiments.
  • To leverage genetic variation across mouse strains for a robust interference assessment.
  • To provide a tool for improving the accuracy of protein abundance profiling.

Main Methods:

  • Development of a strain-specific peptide (SSP) interference reference sample using TMTpro labeling.
  • Utilizing genetic differences in eight mouse strains to identify interference.
  • Quantification of interference using the interference-free index (IFI).
  • Evaluation of interference across three data acquisition methods and ion mobility-based gas-phase fractionation.

Main Results:

  • The SSP effectively identifies and quantifies peptide interference in TMT-based proteomics.
  • Ion mobility-based gas-phase fractionation significantly improves the IFI.
  • Demonstrated the utility of SSP across different data acquisition strategies.
  • Developed an online viewer for data visualization and IFI calculation.

Conclusions:

  • The SSP is a valuable quality control for isobaric tag-based proteomics.
  • This resource aids in the optimization of experimental design and data analysis.
  • Improved accuracy in protein abundance profiling is achievable with the SSP and ion mobility techniques.