Synchronized Stepped Frequency Modulation for Multiplexed Ion Mobility Measurements

Abstract

Implementation of frequency-encoded multiplexing for ion mobility spectrometry (e.g., Fourier transform ion mobility spectrometry (FT-IMS)) has facilitated the direct coupling of drift tube ion mobility instrumentation with ion-trap mass analyzers despite their duty cycle mismatch. Traditionally, FT-IMS experiments have been carried out to utilize continuous linear frequency sweeps that are independent of the scan rate of the ion-trap mass analyzer, thus creating a situation where multiple frequencies are swept over two sequential mass scans. This in turn creates a degree of ambiguity in which the ion current derived from a single modulation frequency cannot be assigned to a single data point in the frequency-modulated signal. In an effort to eliminate this ambiguity, this work describes a discrete stepwise function to modulate the ion gates of the IMS while synchronization between the generated frequencies and the scan rate of the linear ion trap is achieved. While the number of individual frequencies used in the stepped frequency sweeps is less than in continuous linear modulation experiments, there is no loss in performance and high levels of precision are maintained across differing combinations of terminal frequencies and scan lengths. Furthermore, the frequency-scan synchronization enables further data-processing techniques such as linear averaging of the frequency modulated signal to drastically improve signal-to-noise ratio for both high and low intensity analytes.

Considerations for Generating Frequency Modulation Waveforms for Fourier Transform-Ion Mobility Experiments

“…Regardless of how aliasing manifests in the final reconstructed arrival time distribution, preventing such experimental error will require an initial evaluation of instrumental parameters (e.g., mass spectrometer scan rate) to create appropriate frequency sweeps with desired maximum drift times. Once acceptable sweep rates are determined, optimization in terms of length and frequency coverage may be performed in order to optimize for signal-to-noise ratio and resolving power. , Assuming a priori knowledge of the compounds used, it is easy to distinguish whether correct mobilities have been recovered and apply straightforward corrections to the experimental method. If this is not the case and an experimentalist is working with unknowns, it may prove to be useful to introduce large internal standards where the standard is the lowest-mobility compound in solution.…”

“…These lengths were chosen in order to successively alias each analyte as shown in Figure . The sweep files were loaded onto an Arduino-compatible microcontroller (Wio Terminal, Seeedstudio) with firmware developed in-house . The microcontroller was interfaced with an Analog AD9850 waveform generator in order to drive the ion gates.…”

“…The sweep files were loaded onto an Arduino-compatible microcontroller (Wio Terminal, Seeedstudio) with firmware developed in-house. 28 The microcontroller was interfaced with an Analog AD9850 waveform generator in order to drive the ion gates. Synchronization between frequency iteration and MS scans was carried out using the TTL (transistor-transistor logic) output from the LTQ and used as a trigger for the microcontrollers to iterate onto the next frequency.…”

“…In continuous modulation experiments, the sweeping of frequencies is independent of the mass spectrometer’s scan rate where only the start and the end of an experiment are synchronized. ,, Another recently described method, discrete stepwise frequency modulation, requires all mass scans and the iteration of frequencies to be synchronized throughout the entirety of the experiment. This is done in order to unambiguously assign ion-current intensity to single frequencies within the sweep . Due to the increased precision of the stepped frequency modulation approach, work and discussions within this paper will be within the context of the stepped modulation approach.…”

“…This is done in order to unambiguously assign ion-current intensity to single frequencies within the sweep. 20 Due to the increased precision of the stepped frequency modulation approach, work and discussions within this paper will be within the context of the stepped modulation approach. However, the principles discussed here also apply to continuous frequency sweeps, albeit to a lesser extent as ion-current derived from unsynchronized modulation cannot be precisely defined as per eqs 1 and 2.…”