Four-dimensional proteomics analysis of human cerebrospinal fluid with trapped ion mobility spectrometry using PASEF

Dong Gi Mun, Rohit Budhraja, Firdous A. Bhat, Roman M. Zenka, Kenneth L. Johnson, Abhay Moghekar, Akhilesh Pandey

Research output: Contribution to journalArticlepeer-review


A quadrupole time-of-flight mass spectrometer coupled with a trapped ion mobility spectrometry (timsTOF) operated in parallel accumulation-serial fragmentation (PASEF) mode has recently emerged as a platform capable of providing four-dimensional (4D) features comprising of elution time, collision cross section (CCS), mass-to-charge ratio, and intensity of peptides. The PASEF mode provides ∼100% ion sampling efficiency both in data-dependent acquisition (DDA) and data-independent acquisition (DIA) modes without sacrificing sensitivity. In addition, targeted measurements using PASEF integrated parallel reaction monitoring (PRM) mode have also been described. However, only limited number of studies have used timsTOF for analysis of clinical samples. Although Orbitrap mass spectrometers have been used for biomarker discovery from cerebrospinal fluid (CSF) in a variety of neurological diseases, these Orbitrap-derived datasets cannot readily be applied for driving experiments on timsTOF mass spectrometers. We generated a catalog of peptides and proteins in human CSF in DDA mode on a timsTOF mass spectrometer and used these data to build a spectral library. This strategy allowed us to use elution times and ion mobility values from the spectral library to design PRM experiments for quantifying previously discovered biomarkers from CSF samples in Alzheimer's disease. When the same samples were analyzed using a DIA approach combined with a spectral library search, a higher number of proteins were identified than in a library-free approach. Overall, we have established a spectral library of CSF as a resource and demonstrated its utility for PRM and DIA studies, which should facilitate studies of neurological disorders.

Original languageEnglish (US)
Article number2200507
Issue number10
StatePublished - May 2023


  • Alzheimer's disease
  • CSF
  • diaPASEF
  • prm-PASEF

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology


Dive into the research topics of 'Four-dimensional proteomics analysis of human cerebrospinal fluid with trapped ion mobility spectrometry using PASEF'. Together they form a unique fingerprint.

Cite this