Differential or field asymmetric waveform ion mobility spectrometry (FAIMS), which filters ions in gases by the difference between mobilities in strong and weak electric fields, has emerged as a powerful analytical separation. Its major attraction is substantial orthogonality to other separations (including conventional IMS based on the absolute ion mobility) and mass spectrometry (MS), which renders FAIMS/MS an impressive platform for a broad range of applications. Over the last several years, the resolving power and peak capacity of FAIMS (and thus of FAIMS/MS) have been increased by an order of magnitude through the use of stronger electric fields, new buffer gas compositions, and extended ion residence times. With hydrogen-rich gas mixtures, the resolving power for peptides and proteins has reached 500. Such high-resolution FAIMS has proven capable of distinguishing very similar species, for example isotopomers and large peptides with variably localized post-translational modifications (PTMs).Previously, high-resolution FAIMS has been coupled to single-stage MS analyses of moderate resolution. Here, we report integration of high-resolution FAIMS to an Orbitrap Velos MS/MS instrument, which features the resolving power up to 100,000 and enables diverse fragmentation modes comprising regular collision-induced dissociation (CID), high-energy CID (HCD), and electron transfer dissociation (ETD). We will discuss the operational challenges of this system, arising from high background pressure caused by the aspiration of hydrogen from FAIMS buffer through the MS inlet, and means to overcome them. Exemplary bionalytical applications, involving a priori PTM localization for peptides and fragmentation of proteins via CID and ETD, will be presented.

High-resolution differential ion mobility separations with tandem mass spectrometry using Orbitrap and its applications

246th National Meeting (2013)

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