Removal of Interference MS/MS Spectra for Accurate Quantification in Isobaric Tag-Based Proteomics

Area of Science:

• Proteomics
• Mass Spectrometry
• Quantitative Biology

Background:

• Mass spectrometry (MS) enables comprehensive proteome analysis, but accurate protein quantification remains a challenge.
• Isobaric tags for relative and absolute quantification (iTRAQ) offers high precision but suffers from reduced accuracy due to interference peptides.
• Interference peptides coelute and coisolate with target peptides, compromising quantitative results in iTRAQ experiments.

Purpose of the Study:

• To develop a novel method, removal of interference mixture MS/MS spectra (RiMS), to enhance the quantification accuracy of isobaric tag approaches.
• To address the issue of decreased accuracy in iTRAQ quantification caused by interfering peptides.

Main Methods:

• Developed RiMS to identify and remove interference from MS/MS spectra.
• Interference detection is based on analyzing the overlap in elution times of scanned precursor ions.
• RiMS does not require specialized MS instruments (e.g., MS3) or additional ion separation techniques.

Main Results:

• Implementing RiMS led to an 11% decrease in protein identification but significantly improved quantification accuracy.
• The method successfully identified differential protein expressions between human-induced pluripotent stem cells and human dermal fibroblasts.
• RiMS demonstrated its utility in quantitative comparisons reflecting distinct cellular characteristics.

Conclusions:

• RiMS is an effective strategy to improve the accuracy of quantitative proteomics using isobaric tagging.
• The method offers a practical solution for enhancing data reliability without necessitating advanced instrumentation.
• RiMS facilitates accurate quantitative comparisons of complex biological samples, such as stem cells and fibroblasts.