A Modular Variable Temperature FT-IMS Instrument Optimized for Gas-Phase Ion Chemistry Applications.

The latest iteration of modular, open-source rolled ion mobility spectrometers was characterized and tailored for heated ion chemistry experiments. Because the nature of ion-neutral interactions is innately linked to the temperature of the drift cell, heated IMS experiments explicitly probe the fundamental characteristics of these collisions. While classic mobility experiments examine ions through inert buffer gases, doping the drift cell with reactive vapor enables desolvated chemical reactions to be studied. By using materials with minimal outgassing and ensuring the isolation of the drift tube from the surrounding ambient conditions, an open-source drift cell outfitted with heating components enables investigations of chemical reactions as a function of temperature. We show here that elevated temperatures facilitate an increase in deuterium incorporation and allow for hydrogen/deuterium exchanges otherwise unattainable under ambient conditions. While the initial fast exchanges get faster as temperature is increased, the slow rate which rises from the kinetic nonlinearity though to be attributed to ion-neutral clustering, remains constant with no change in mobility shifts. Additionally, we show the analytical merit of multiplexing mobility data by comparing the performance of traditional signal-averaging and FT-IMS modes.

Nonlinear Frequency Modulation for Fourier Transform Ion Mobility Mass Spectrometry Improves Experimental Efficiency.

Through optimization of terminal frequencies and effective sampling rates, we have developed nonlinear sawtooth-shaped frequency sweeps for efficient Fourier transform ion mobility mass spectrometry (FT-IM-MS) experiments. This is in contrast to conventional FT-IM-MS experiments where ion gates are modulated according to a linear frequency sweep. Linear frequency sweeps are effective but can be hindered by the amount of useful signal obtained using a single sweep over a large frequency range imposed by ion gating inefficiencies, particularly small ion packets, and gate depletion. These negative factors are direct consequences of the inherently low gate pulse widths of high-frequency ion gating events, placing an upper bound on FT-IM-MS performance. Here, we report alternative ion modulation strategies. Sawtooth frequency sweeps may be constructed for the purpose of either extending high-SNR transients or conducting efficient signal-averaging experiments for low-SNR transients. The data obtained using this approach show high-SNR signals for a set of low-mass tetraalkylammonium salts (<1000 m/z) where resolving powers in excess of 500 are achieved. Data for low-SNR obtained for multimeric protein complexes streptavidin (53 kDa) and GroEL (800 kDa) also reveal large increases in the signal-to-noise ratio for reconstructed arrival time distributions.

A Comparison of the Performance of Modular Standalone Do-It-Yourself Ion Mobility Spectrometry Systems.

As the spectrum of ion mobility spectrometry (IMS) applications expands and more experimental configurations are developed, identifying the correct platform for an experimental campaign becomes more challenging for researchers. Additionally, metrics that compare performance (Rp, for example) often have nuanced differences in definition between platforms that render direct comparisons difficult. Here we present a comparison of three do-it-yourself (DIY) drift tubes that are relatively low cost and easy to construct, where the performance of each is evaluated based on three different metrics: resolving power, the ideality of resolving powers, and accuracy/precision of K0 values. The standard PCBIMS design developed by Reinecke and Clowers (Reinecke, T.; Clowers, B. H. HardwareX 2018, 4, e00030) provided the highest resolving power (>90) and the highest ideality of resolving power ratios (>90% at best) of the three systems. However, the flexible tube (FlexIMS) construction as described by Smith et al. (Smith, B. L. Anal. Chem. 2020, 92 (13), 9104-9112) exhibited the highest degree of precision of K0 values (relative standard deviation of <0.42%). Depending on the application, the drift tube variants presented and evaluated here offer a low-cost alternative to commercial drift-tube systems with levels of performance that approach theoretical maxima.

FTflow: An Open-Source Python GUI for FT-IM-MS Experiments.

As part of a larger effort to aid in seamless integration of Fourier-based multiplexed ion mobility with a range mass analyzers, we have developed an all-in-one graphical user interface tool for FT-IM-MS data analysis that runs directly within a web browser. This tool, FTflow, accepts mzML files and displays necessary information such as mass spectra and extracted ion chromatograms in order to reconstruct arrival time distributions. It also extracts the corresponding mobility-related information (e.g., Ko and CCS) for each of the target ion populations. Furthermore, input fields for experimental conditions are clearly laid out for users and ease-of-use. With flexibility in mind, the processing scripts and GUI interface are written entirely in Python and allows users the option to modify source code to fit their specific needs. While the intention for this tool is to be a starting point for exploratory analysis of FT-IM-MS data, it has the capability to be adapted for use in more automated data processing pipelines through direct access of core processing routines.

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

By casting the information regarding an ion population's mobility in the frequency domain, the coupling of time-dispersive ion mobility techniques is now imminently compatible with slower mass analyzers such as ion traps. Recent reports have detailed the continued progress toward maximizing the efficiency of the Fourier transform ion mobility-mass spectrometry (FT-IM-MS) experiments, but few reports have outlined the intersection between the practical considerations of implementation against the theoretical limits imposed by traditional signal processing techniques. One of the important concerns for Fourier-based multiplexing experiments is avoiding signal aliasing as a product of undersampled signals that may occur during data acquisition. In addition to traditional considerations such as detector sampling frequency, the limitations (i.e., maximum measurable drift time) imposed by experimental mass scan duration and the frequency sweep used for ion gate modulation must also be assessed. This work aims to connect the fundamental underpinnings of FT-IM-MS experiments and the associated experimental parameters that are encountered when coupling the comparatively fast separations in the mobility domain with the slower m/z scanning common for ion-trap mass analyzers. In addition to stating the relevant theory that applies to the FT-IM-MS experiment, this report highlights how aliased signals will manifest post Fourier transform in reconstructed arrival time distributions and calculated mobilities.

Synchronized Stepped Frequency Modulation for Multiplexed Ion Mobility Measurements.

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.