A systematic optimization of diagonal-PASEF acquisitions for low-to-medium input on the timsTOF HT mass spectrometer

Liquid chromatography mass spectrometry (LC-MS) methods aim to yield high peptide and protein identifications while retaining quantitative accuracy and precision. To increase the analytical performance of the timsTOF mass spectrometers, researchers have recently proposed diagonal dia-PASEF acquisition methods termed as synchro- or midia-PASEF which attempt to acquire the ion cloud more efficiently (Skowronek, 2023; Distler, 2023). These methods operate by seamlessly and continuously following the observed diagonal shape of the precursor ion density. While such methods promise to cover the observed ion-cloud more efficiently, their utility is currently still hampered by a lack of systematic method evaluation. We therefore set out to optimize the diagonal-PASEF acquisitions on a timsTOF-HT mass spectrometer for different applications.