Synchro-PASEF Allows Precursor-Specific Fragment Ion Extraction and Interference Removal in Data-Independent...

Area of Science:

• Mass Spectrometry
• Proteomics
• Analytical Chemistry

Background:

• Data-independent acquisition (DIA) is widely used in mass spectrometry-based proteomics.
• DIA enables simultaneous acquisition of fragment spectra for all precursors.
• Reconstructing precursor-fragment relationships in DIA is challenging due to convoluted spectra.

Purpose of the Study:

• Introduce synchro-PASEF, a new scan mode for trapped ion mobility spectrometry-parallel accumulation-serial fragmentation (TIMS-PASEF).
• Improve the reconstruction of precursor-fragment relationships in DIA.
• Enhance the accuracy of proteomic identification and quantification.

Main Methods:

• Developed synchro-PASEF, a scan mode that follows ion cloud shapes in ion mobility and mass dimensions.
• Consecutive quadrupole selection windows move synchronously, slicing peptide precursors.
• Separates fragment ion signals into adjacent scans for precise precursor-fragment definition.

Main Results:

• Synchro-PASEF increases detected fragment ion current several-fold at sub-second cycle times.
• Precisely defines precursor-fragment relationships in ion mobility and mass dimensions.
• Effectively deconvolutes DIA fragment space and enhances quantification specificity.

Conclusions:

• Synchro-PASEF precisely defines precursor-fragment relationships, overcoming DIA challenges.
• The method offers improved specificity and accuracy for proteomic analyses.
• Synchro-PASEF has significant potential for advancing quantitative proteomics.