Improving Ion Mobility Mass Spectrometry of Proteins through Tristate Gating and Optimization of Multiplexing Parameters

Abstract

Coupling drift tube ion mobility (IM) to Fourier transform mass spectrometry (FT-MS) affords the opportunity for gas-phase separation of ions based on size and conformation with high-resolution mass analysis. However, combining IM and FT-MS is challenging because ions exit the drift tube on a much faster time scale than the rate of mass analysis. Fourier transform (FT) and Hadamard transform multiplexing methods have been implemented to overcome the duty-cycle mismatch, offering new avenues for obtaining high-resolution, high-mass-accuracy analysis of mobility-selected ions. The gating methods used to integrate the drift tube with the FT mass analyzer discriminate against the transmission of large, low-mobility ions owing to the well-known gate depletion effect. Tristate gating strategies have been shown to increase ion transmission for drift tube IM-FT-MS systems through implementation of dual ion gating, controlling the quantity and timing of ions through the drift tube to reduce losses of slow-moving ions. Here we present an optimized set of multiplexing parameters for tristate gating ion mobility of several proteins on an Orbitrap mass spectrometer and further report parameters for increased ion transmission and mobility resolution as well as decreased experimental times from 15 min down to 30 s. On average, peak intensities in the arrival time distributions (ATDs) for ubiquitin increased 2.1× on average, while those of myoglobin increased by 1.5× with a resolving power increase on average of 11%.

Electrospray Ion Mobility Spectrometer Based on Flexible Printed-Circuit Board Electrodes with Improved Resolving Power

“…The use of tristate injection has been suggested in order to minimize a discrimination of ions during the injection. The working principle is described elsewere. ,, To investigate the possibility of tristate injection on the new instrument, the pulsing regime was implemented by combining two pulser circuitries as reported by the Clowers’ group . Peak ion currents and resolving powers obtained for the lower-mobility ions, namely, T8, T10, and T12, were compared and are tabulated in Table .…”

“… 32 , 41 , 42 To investigate the possibility of tristate injection on the new instrument, the pulsing regime was implemented by combining two pulser circuitries as reported by the Clowers’ group. 32 Peak ion currents and resolving powers obtained for the lower-mobility ions, namely, T8, T10, and T12, were compared and are tabulated in Table 4 . These compounds were introduced separately to avoid any possible bias in the measurements by ion suppression effects affecting ESI.…”

“…These are battery operated, floated to the high voltage at the gate, and triggered via an optical fiber to maintain the isolation. The circuitries are given in Garcia et al 31 Two of these were wired in series, and the connection point was attached to the middle grid, as described by Butalewicz et al 32 This allowed the control of the electric field at the gate in a two-state or tristate fashion. Triggering was achieved with an Analog Discovery 2 unit (Digilent, Pullman, WA).…”

Instrumentation at the Leading Edge of Proteomics

“…Separation quality is affected by drift tube length, which is limited by the need for high drift voltages across the drift region. Canonically, the speed of ion mobility separations, in microseconds to milliseconds, has driven the dominance of Q-TOF mass spectrometers as mass analyzers for ion mobility systems, but ongoing work has also coupled DTIMS to FTICR and Orbitrap systems. − The development of Fourier transform IMS, modular DTIMS instrumentation, and cheaper alternative power supplies is poised to enable more accessible MS instrumentation and development, especially for IMS. − …”

Ultra-Fast Ion Mobility Spectrometer for High-Throughput Chromatography

“…In this context, the choice of the ion shutter is decisive for the reduction of the initial ion packet width. In the present work, we use a tristate ion shutter which exhibits no ion discrimination of less mobile ions even for shortest shutter opening times. − For most other ion shutters, ion discrimination creates a limit for the lowest initial ion packet width that is still practically useable due to increasing loss of sensitivity with decreasing shutter opening times.…”