Cyclable Variable Path Length Multilevel Structures for Lossless Ion Manipulations (SLIM) Platform for Enhanced Ion Mobility Separations.

Ion mobility-mass spectrometry (IMS-MS) is used to analyze complex samples and provide structural information on unknown compounds. As the complexity of samples increases, there is a need to improve the resolution of IMS-MS instruments to increase the rate of molecular identification. This work evaluated a cyclable and variable path length (and hence resolving power) multilevel Structures for Lossless Ion Manipulations (SLIM) platform to achieve a higher resolving power than what was previously possible. This new multilevel SLIM platform has eight separation levels connected by ion escalators, yielding a total path length of ∼88 m (∼11 m per level). Our new multilevel SLIM can also be operated in an "ion cycling" mode by utilizing a set of return ion escalators that transport ions from the eighth level back to the first, allowing even extendable path lengths (and higher IMS resolution). The platform has been improved to enhance ion transmission and IMS separation quality by reducing the spacing between SLIM boards. The board thickness was reduced to minimize the ions' escalator residence time. Compared to the previous generation, the new multilevel SLIM demonstrated better transmission for a set of phosphazene ions, especially for the low-mobility ions. For example, the transmission of m/z 2834 ions was improved by a factor of ∼3 in the new multilevel SLIM. The new multilevel SLIM achieved 49% better resolving powers for GRGDS1+ ions in 4 levels than our previous 4-level SLIM. The collision cross-section-based resolving power of the SLIM platform was tested using a pair of reverse sequence peptides (SDGRG1+, GRGDS1+). We achieved 1100 resolving power using 88 m of path length (i.e., 8 levels) and 1400 following an additional pass through the eight levels. Further evaluation of the multilevel SLIM demonstrated enhanced separation for positively and negatively charged brain total lipid extract samples. The new multilevel SLIM enables a tunable high resolving power for a wide range of ion mobilities and improved transmission for low-mobility ions.

Optimization of a Digital Mass Filter for the Isolation of Intact Protein Complexes in Stability Zone 1,1.

Digital mass filters are advantageous for the analysis of large molecules due to the ability to perform ion isolation of high-m/z ions without the generation of very high radio frequency (RF) and DC voltages. Experimentally determined Mathieu stability diagrams of stability zone 1,1 for capacitively coupled digital waveforms show a voltage offset between the quadrupole rod pairs is introduced by the capacitors which is dependent on the voltage magnitude of the waveform and the duty cycle. This changes the ion's a value from a = 0 to a < 0. These effects are illustrated for isolation for single-charge states for various protein complexes up to 800 kDa (GroEL) for stability zone 1,1. Isolation resolving power (m/Δm) of approximately 280 was achieved for an ion of m/z 12,315 (z = 65+ for 800.5 kDa GroEL D398A), which corresponds to an m/z window of 44.

A Dual-Gated Structures for Lossless Ion Manipulations-Ion Mobility Orbitrap Mass Spectrometry Platform for Combined Ultra-High-Resolution Molecular Analysis.

High-resolution ion mobility spectrometry-mass spectrometry (HR-IMS-MS) instruments have enormously advanced the ability to characterize complex biological mixtures. Unfortunately, HR-IMS and HR-MS measurements are typically performed independently due to mismatches in analysis time scales. Here, we overcome this limitation by using a dual-gated ion injection approach to couple an 11 m path length structures for lossless ion manipulations (SLIM) module to a Q-Exactive Plus Orbitrap MS platform. The dual-gate setup was implemented by placing one ion gate before the SLIM module and a second ion gate after the module. The dual-gated ion injection approach allowed the new SLIM-Orbitrap platform to simultaneously perform an 11 m SLIM separation, Orbitrap mass analysis using the highest selectable mass resolution setting (up to 140 k), and high-energy collision-induced dissociation (HCD) in ∼25 min over an m/z range of ∼1500 amu. The SLIM-Orbitrap platform was initially characterized using a mixture of standard phosphazene cations and demonstrated an average SLIM CCS resolving power (RpCCS) of ∼218 and an SLIM peak capacity of ∼156, while simultaneously obtaining high mass resolutions. SLIM-Orbitrap analysis with fragmentation was then performed on mixtures of standard peptides and two reverse peptides (SDGRG1+, GRGDS1+, and RpCCS = 305) to demonstrate the utility of combined HR-IMS-MS/MS measurements for peptide identification. Our new HR-IMS-MS/MS capability was further demonstrated by analyzing a complex lipid mixture and showcasing SLIM separations on isobaric lipids. This new SLIM-Orbitrap platform demonstrates a critical new capability for proteomics and lipidomics applications, and the high-resolution multimodal data obtained using this system establish the foundation for reference-free identification of unknown ion structures.

Top/Middle-Down Characterization of α-Synuclein Glycoforms.

α-Synuclein is an intrinsically disordered protein that plays a critical role in the pathogenesis of neurodegenerative disorders, such as Parkinson's disease. Proteomics studies of human brain samples have associated the modification of the O-linked N-acetyl-glucosamine (O-GlcNAc) to several synucleinopathies; in particular, the position of the O-GlcNAc can regulate protein aggregation and subsequent cell toxicity. There is a need for site specific O-GlcNAc α-synuclein screening tools to direct better therapeutic strategies. In the present work, for the first time, the potential of fast, high-resolution trapped ion mobility spectrometry (TIMS) preseparation in tandem with mass spectrometry assisted by an electromagnetostatic (EMS) cell, capable of electron capture dissociation (ECD), and ultraviolet photodissociation (213 nm UVPD) is illustrated for the characterization of α-synuclein positional glycoforms: T72, T75, T81, and S87 modified with a single O-GlcNAc. Top-down 213 nm UVPD and ECD MS/MS experiments of the intact proteoforms showed specific product ions for each α-synuclein glycoforms associated with the O-GlcNAc position with a sequence coverage of ∼68 and ∼82%, respectively. TIMS-MS profiles of α-synuclein and the four glycoforms exhibited large structural heterogeneity and signature patterns across the 8+-15+ charge state distribution; however, while the α-synuclein positional glycoforms showed signature mobility profiles, they were only partially separated in the mobility domain. Moreover, a middle-down approach based on the Val40-Phe94 (55 residues) chymotrypsin proteolytic product using tandem TIMS-q-ECD-TOF MS/MS permitted the separation of the parent positional isomeric glycoforms. The ECD fragmentation of the ion mobility and m/z separated isomeric Val40-Phe94 proteolytic peptides with single O-GlcNAc in the T72, T75, T81, and S87 positions provided the O-GlcNAc confirmation and positional assignment with a sequence coverage of ∼80%. This method enables the high-throughput screening of positional glycoforms and further enhances the structural mass spectrometry toolbox with fast, high-resolution mobility separations and 213 nm UVPD and ECD fragmentation capabilities.

A planar quadrupole device for transmitting and trapping ions in high vacuum.

RATIONALE: Hybrid mass spectrometers combine multiple mass analyzers to achieve optimal performance in terms of tandem mass spectrometry, high mass resolving power, and mass measurement accuracy for studying highly complex samples. As a result, the need for transport, trapping, and control of ion kinetic energies is critical for the successful integration of multiple mass analyzers and hybrid instrument operation. In addition, transportation of ion populations between two physically distinct locations can result in time-of-flight (TOF) discrimination against ions with widely disparate m/z values, compromising full mass spectral performance. In this work, we demonstrated a new ion guide, referred to as a planar quadrupole (PQ) ion guide, composed of two parallel printed circuit boards (PCB) that allow radiofrequency (RF) and direct current (DC) voltages to be combined to enable both axial transport and trapping of ion populations in the ultrahigh vacuum region of the mass spectrometer. As compared with a conventional multipole ion guide, the PQ ion guide showed comparable performance in ion m/z values, signal-to-noise, and intensity and effectively reduced mass discrimination caused by TOF effects. METHODS: A PQ device was developed with two PCBs and simulated with SIMION 8.1. Electrospray ionization and Fourier transform ion cyclotron resonance mass spectrometry instrumentation were used for the testing of PQ performance. RESULTS: .In this work, we demonstrated a planar quadrupole (PQ) ion guide composed of two parallel PCB plates. The PQ enables both axial ion transport and trapping of ion populations throughout the ion transfer process from a LTQ to an ICR cell. As compared with a conventional multipole ion guide, the PQ showed comparable ion transmission efficiency and effectively reduced mass discrimination caused by TOF effects. CONCLUSIONS: The PQ is a simple design that can be implemented for ion transmission and trapping on virtually any mass spectrometer.

Application of frequency multiple FT-ICR-MS signal acquisition for improved proteome research.

Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) coupled with liquid chromatography (LC) is a powerful combination useful in many research areas due to the utility of high mass resolving power and mass measurement accuracy for studying highly complex samples. Ideally, every analyte in a complex sample can be subjected to accurate mass MS/MS analysis to aid in identification. FT-ICR MS can provide high mass resolving power and mass accuracy at the cost of long data acquisition periods, reducing the number of spectra that can be acquired per unit time. Frequency multiple signal acquisition has long been realized as an attractive method to obtain high mass resolving power and mass accuracy with shorter data acquisition periods. However, one of the limitations associated with frequency multiple signal acquisition is reduced signal intensity as compared to a traditional dipole detector. In this study, we demonstrated the use of a novel ICR cell to improve frequency multiple signal intensity and investigated the potential use of frequency multiple acquisition for proteome measurements. This novel ICR cell containing both dipole and frequency multiple detection electrodes was installed on a 7T FT-ICR MS coupled to an LC system. Tryptic digests of HeLa cell lysates were analyzed using dipole and frequency multiple detectors by holding either the mass resolving power or signal acquisition time constant. Compared to dipole detection, second frequency multiple detection yielded 36% or 45% more unique identified peptides from HeLa cell lysates at twice the scan rate or twice the mass resolving power, respectively. These results indicate that frequency multiple signal acquisition with either the same resolving power or the same signal acquisition duration as used with dipole signals can produce a significant increase in the number of peptides identified in complex proteome samples.

Ultra-High-Resolution Ion Mobility Separations Over Extended Path Lengths and Mobility Ranges Achieved using a Multilevel Structures for Lossless Ion Manipulations Module.

Over the past few years, structures for lossless ion manipulations (SLIM) have used traveling waves (TWs) to move ions over long serpentine paths that can be further lengthened by routing the ions through multiple passages of the same path. Such SLIM "multipass" separations provide unprecedentedly high ion mobility resolving powers but are ultimately limited in their ion mobility range because of the range of mobilities spanned in a single pass; that is, higher mobility ions ultimately "overtake" and "lap" lower mobility ions that have experienced fewer passes, convoluting their arrival time distribution at the detector. To achieve ultrahigh resolution separations over broader mobility ranges, we have developed a new multilevel SLIM possessing multiple stacked serpentine paths. Ions are transferred between SLIM levels through apertures (or ion escalators) in the SLIM surfaces. The initial multilevel SLIM module incorporates four levels and three interlevel ion escalator passages, providing a total path length of 43.2 m. Using the full path length and helium buffer gas, high resolution separations were achieved for Agilent tuning mixture phosphazene ions over a broad mobility range (K0 ≈ 3.0 to 1.2 cm2/(V*s)). High sensitivity was achieved using "in-SLIM" ion accumulation over an extended trapping region of the first SLIM level. High transmission efficiency of ions over a broad mobility range (e.g., K0 ≈ 3.0 to 1.67 cm2/(V*s)) was achieved, with transmission efficiency rolling off for the lower mobility ions (e.g., K0 ≈ 1.2 cm2/(V*s)). Resolving powers of up to ∼560 were achieved using all four ion levels to separate reverse peptides (SDGRG1+ and GRGDS1+). A complex mixture of phosphopeptides showed similar coverage could be achieved using one or all four SLIM levels, and doubly charged phosphosite isomers not significantly separated using one SLIM level were well resolved when four levels were used. The new multilevel SLIM technology thus enables wider mobility range ultrahigh-resolution ion mobility separations and expands on the ability of SLIM to obtain improved separations of complex mixtures with high sensitivity.

Design and Performance of a Dual-Polarity Instrument for Ion Soft Landing.

A new apparatus for ion soft landing research was developed and is reported in this contribution. The instrument includes a dual polarity high-flux electrospray ionization (ESI) interface, a tandem electrodynamic ion funnel system, a collisional flatapole, a quadrupole mass filter, and a focusing lens. The instrument enables production of ionic layers by soft landing of mass-selected ions onto surfaces with balanced or imbalanced charge conditions using either layer-by-layer (LBL) or fast polarity switching modes. We present the first evidence of using weakly coordinating stable anions to protect the ionizing protons of soft-landed cations on the surface. The observed proton retention is particularly efficient when fast polarity switching of anions and cations is employed to deposit small quantities of ions in short deposition segments. Furthermore, we observe more efficient charge retention and better ionic complexation in a charge-balanced layer prepared by fast polarity switching deposition. These findings open up new opportunities for the fabrication of novel ionic assemblies using well-defined gaseous ions as building blocks.

Differential Ion Mobility Separations/Mass Spectrometry with High Resolution in Both Dimensions.

Strong orthogonality to mass spectrometry makes differential ion mobility spectrometry (FAIMS) a powerful tool for isomer separations. However, high FAIMS resolution has been achieved overall only with buffers rich in He or H2. That obstructed coupling to Fourier transform mass spectrometers operating under ultrahigh vacuum, but exceptional m/ z resolution and accuracy of FTMS are indispensable for frontline biological and environmental applications. By raising the waveform amplitude to 6 kV, we enabled high FAIMS resolution using solely N2 and thus straightforward integration with any MS platform: here Orbitrap XL with the electron transfer dissociation (ETD) option. The initial evaluation for complete histone tails (50 residues) with diverse post-translational modifications on alternative sites demonstrates a broad capability to separate and confidently identify the PTM localization variants in the middle-down range.

Traveling-Wave-Based Electrodynamic Switch for Concurrent Dual-Polarity Ion Manipulations in Structures for Lossless Ion Manipulations.

We describe the development of a dual-polarity traveling-wave (TW) structures for lossless ion manipulations (SLIM) ion mobility spectrometry (IMS) device capable of switching both positive and negative ions that are traveling simultaneously along the same path to different regions of the SLIM. Through simulations, the routing efficiency of the SLIM TW switch was compared to a SLIM direct-current-based (DC) switch developed previously for IMS-MS. We also report on the initial experimental evaluation of a dual-polarity SLIM platform, which uses the TW-based ion switch to achieve higher resolution multipass serpentine ultralong path with extended routing (SUPER) IMS separations. Overall, these results show that the dual-polarity TW switch is not only as effective as DC switching in terms of routing efficiency but also is agnostic to the polarity of the ions being routed.

High-Resolution Differential Ion Mobility Separations/Orbitrap Mass Spectrometry without Buffer Gas Limitations.

Strong orthogonality between differential ion mobility spectrometry (FAIMS) and mass spectrometry (MS) makes their hybrid a powerful approach to separate isomers and isobars. Harnessing that power depends on high resolution in both dimensions. The ultimate mass resolution and accuracy are delivered by Fourier Transform MS increasingly realized in Orbitrap MS, whereas FAIMS resolution is generally maximized by buffers rich in He or H2 that elevate ion mobility and lead to prominent non-Blanc effects. However, turbomolecular pumps have lower efficiency for light gas molecules and their flow from the FAIMS stage complicates maintaining the ultrahigh vacuum (UHV) needed for Orbitrap operation. Here we address this challenge via two hardware modifications: (i) a differential pumping step between FAIMS and MS stages and (ii) reconfiguration of vacuum lines to isolate pumping of the high vacuum (HV) region. Either greatly ameliorates the pressure increases upon He or H2 aspiration. This development enables free optimization of FAIMS carrier gas without concerns about MS performance, maximizing the utility and flexibility of FAIMS/MS platforms.

Parallel detection in a single ICR cell: Spectral averaging and improved S/N without increased acquisition time.

Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) is well-renowned for its ultrahigh resolving power and mass measurement accuracy. As with other types of analytical instrumentation, achievable signal-to-noise ratio (S/N) is an important analytical figure of merit with FTICR-MS. S/N can be improved with higher magnetic fields and longer time-domain signal acquisition periods. However, serial signal averaging of spectra or time-domain signals acquired with multiple ion populations is most commonly used to improve S/N. On the other hand, serial acquisition and averaging of multiple scans significantly increases required data acquisition time and is often incompatible with on-line chromatographic separations. In this study, we investigated the potential for increased S/N by averaging 4 spectra that were acquired in parallel with a single ICR cell with 4 pairs of dipole detection electrodes, each with an independent pre-amplifier. This spectral averaging was achieved with no need for multiple ion accumulation events nor multiple, serial excitation and detection events. These efforts demonstrated that parallel signal acquisition with 4 detector electrode pairs produces S/N 1.76-fold higher than that from a single detection electrode pair. With parallel detection, improved S/N was achieved with no observable loss in resolving power (100,000) as compared with that from a single detection electrode pair. These results demonstrate that parallel detection of multiple induced image current signals with multiple preamplifiers exists as a viable option for future instrumentation to increase achievable S/N and sensitivity. This approach may have general utility especially where conventional serial signal averaging is impractical.

Design and Performance of a Novel Interface for Combined Matrix-Assisted Laser Desorption Ionization at Elevated Pressure and Electrospray Ionization with Orbitrap Mass Spectrometry.

Matrix-Assisted Laser Desorption Ionization, MALDI, has been increasingly used in a variety of biomedical applications, including tissue imaging of clinical tissue samples, and in drug discovery and development. These studies strongly depend on the performance of the analytical instrumentation and would drastically benefit from improved sensitivity, reproducibility, and mass/spatial resolution. In this work, we report on a novel combined MALDI/ESI interface, which was coupled to different Orbitrap mass spectrometers (Elite and Q Exactive Plus) and extensively characterized with peptide and protein standards, and in tissue imaging experiments. In our approach, MALDI is performed in the elevated pressure regime (5-8 Torr) at a spatial resolution of 15-30 µm, while ESI-generated ions are injected orthogonally to the interface axis. We have found that introduction of the MALDI-generated ions into an electrodynamic dual-funnel interface results in increased sensitivity characterized by a limit of detection of ∼400 zmol, while providing a mass measurement accuracy of 1 ppm and a mass resolving power of 120 000 in analysis of protein digests. In tissue imaging experiments, the MALDI/ESI interface has been employed in experiments with rat brain sections and was shown to be capable of visualizing and spatially characterizing very low abundance analytes separated only by 20 mDa. Comparison of imaging data has revealed excellent agreement between the MALDI and histological images.

Serpentine Ultralong Path with Extended Routing (SUPER) High Resolution Traveling Wave Ion Mobility-MS using Structures for Lossless Ion Manipulations.

Ion mobility (IM) separations have a broad range of analytical applications, but insufficient resolution often limits their utility. Here, we report on ion mobility separations in a structures for lossless ion manipulations (SLIM) serpentine ultralong path with extended routing (SUPER) traveling wave (TW) ion mobility (IM) module in conjunction with mass spectrometry (MS). Ions were confined in the SLIM by rf fields in conjunction with a DC guard bias, enabling essentially lossless TW transmission over greatly extended paths. The extended routing utilized multiple passes (e.g., ∼1094 m over 81 passes through the 13.5 m serpentine path) and was facilitated by the introduction of a lossless ion switch that allowed ions to be directed to either the MS detector or for another pass through the serpentine separation region, allowing theoretically unlimited IM path lengths. The multipass SUPER IM-MS provided resolution approximately proportional to the square root of the number of passes (or total path length). More than 30-fold higher IM resolution (∼340 vs ∼10) for Agilent tuning mix m/z 622 and 922 ions was achieved for 40 passes compared to commercially available drift tube IM and other TWIM-based platforms. An initial evaluation of the isomeric sugars lacto-N-hexaose and lacto-N-neohexaose showed the isomeric structures to be baseline resolved, and a new conformational feature for lacto-N-neohexaose was revealed after 9 passes. The new SLIM SUPER high resolution TWIM platform has broad utility in conjunction with MS and is expected to enable a broad range of previously challenging or intractable separations.

Parallel Spectral Acquisition with an Ion Cyclotron Resonance Cell Array.

Mass measurement accuracy is a critical analytical figure-of-merit in most areas of mass spectrometry application. However, the time required for acquisition of high-resolution, high mass accuracy data limits many applications and is an aspect under continual pressure for development. Current efforts target implementation of higher electrostatic and magnetic fields because ion oscillatory frequencies increase linearly with field strength. As such, the time required for spectral acquisition of a given resolving power and mass accuracy decreases linearly with increasing fields. Mass spectrometer developments to include multiple high-resolution detectors that can be operated in parallel could further decrease the acquisition time by a factor of n, the number of detectors. Efforts described here resulted in development of an instrument with a set of Fourier transform ion cyclotron resonance (ICR) cells as detectors that constitute the first MS array capable of parallel high-resolution spectral acquisition. ICR cell array systems consisting of three or five cells were constructed with printed circuit boards and installed within a single superconducting magnet and vacuum system. Independent ion populations were injected and trapped within each cell in the array. Upon filling the array, all ions in all cells were simultaneously excited and ICR signals from each cell were independently amplified and recorded in parallel. Presented here are the initial results of successful parallel spectral acquisition, parallel mass spectrometry (MS) and MS/MS measurements, and parallel high-resolution acquisition with the MS array system.

Squeezing of Ion Populations and Peaks in Traveling Wave Ion Mobility Separations and Structures for Lossless Ion Manipulations Using Compression Ratio Ion Mobility Programming.

In this work we report an approach for spatial and temporal gas-phase ion population manipulation, wherein we collapse ion distributions in ion mobility (IM) separations into tighter packets providing higher sensitivity measurements in conjunction with mass spectrometry (MS). We do this for ions moving from a conventional traveling wave (TW)-driven region to a region where the TW is intermittently halted or "stuttered". This approach causes the ion packets spanning a number of TW-created traveling traps (TT) to be redistributed into fewer TT, resulting in spatial compression. The degree of spatial compression is controllable and determined by the ratio of stationary time of the TW in the second region to its moving time. This compression ratio ion mobility programming (CRIMP) approach has been implemented using "structures for lossless ion manipulations" (SLIM) in conjunction with MS. CRIMP with the SLIM-MS platform is shown to provide increased peak intensities, reduced peak widths, and improved signal-to-noise (S/N) ratios with MS detection. CRIMP also provides a foundation for extremely long path length and multipass IM separations in SLIM providing greatly enhanced IM resolution by reducing the detrimental effects of diffusional peak broadening and increasing peak widths.

Achieving High Resolution Ion Mobility Separations Using Traveling Waves in Compact Multiturn Structures for Lossless Ion Manipulations.

We report on ion mobility (IM) separations achievable using traveling waves (TW) in a Structures for Lossless Ion Manipulations (SLIM) module having a 44 cm path length and 16 90° turns. The performance of the TW-SLIM module was evaluated for ion transmission and IM separations with different RF, TW parameters, and SLIM surface gaps in conjunction with mass spectrometry. In this work, TWs were created by the transient and dynamic application of DC potentials. The module demonstrated highly robust performance and, even with 16 closely spaced turns, achieving IM resolution performance and ion transmission comparable to a similar straight path module. We found an IM peak capacity of ∼31 and peak generation rate of 780 s(-1) for TW speeds of ∼80 m/s using the current multi-turn TW-SLIM module. The separations achieved for isomers of peptides and tetrasaccharides were found to be comparable to those from a ∼0.9-m drift tube-based IM-MS platform operated at the same pressure (4 Torr). The combined attributes of flexible design, low voltage requirements and lossless ion transmission through multiple turns for the present TW-SLIM module provides a basis for SLIM devices capable of achieving much greater IM resolution via greatly extended ion path lengths and using compact serpentine designs.

Ultra-High Resolution Ion Mobility Separations Utilizing Traveling Waves in a 13 m Serpentine Path Length Structures for Lossless Ion Manipulations Module.

We report the development and initial evaluation of a 13 m path length Structures for Lossless Manipulations (SLIM) module for achieving high resolution separations using traveling waves (TW) with ion mobility (IM) spectrometry. The TW SLIM module was fabricated using two mirror-image printed circuit boards with appropriately configured RF, DC, and TW electrodes and positioned with a 2.75 mm intersurface gap. Ions were effectively confined in field-generated conduits between the surfaces by RF-generated pseudopotential fields and moved losslessly through a serpentine path including 44 "U" turns using TWs. The ion mobility resolution was characterized at different pressures, gaps between the SLIM surfaces, and TW and RF parameters. After initial optimization, the SLIM IM-MS module provided about 5-fold higher resolution separations than present commercially available drift tube or traveling wave IM-MS platforms. Peak capacity and peak generation rates achieved were 246 and 370 s(-1), respectively, at a TW speed of 148 m/s. The high resolution achieved in the TW SLIM IM-MS enabled, e.g., isomeric sugars (lacto-N-fucopentaose I and lacto-N-fucopentaose II) to be baseline resolved, and peptides from an albumin tryptic digest were much better resolved than with existing commercial IM-MS platforms. The present work also provides a foundation for the development of much higher resolution SLIM devices based upon both considerably longer path lengths and multipass designs.

Mobility-Selected Ion Trapping and Enrichment Using Structures for Lossless Ion Manipulations.

The integration of ion mobility spectrometry (IMS) with mass spectrometry (MS) and the ability to trap ions in IMS-MS measurements is of great importance for performing reactions, accumulating ions, and increasing analytical measurement sensitivity. The development of Structures for Lossless Ion Manipulations (SLIM) offers the potential for ion manipulations in an extended and more effective manner, while opening opportunities for many more complex sequences of manipulations. Here, we demonstrate an ion separation and trapping module and a method based upon SLIM that consists of a linear mobility ion drift region, a switch/tee and a trapping region that allows the isolation and accumulation of mobility-separated species. The operation and optimization of the SLIM switch/tee and trap are described and demonstrated for the enrichment of the low abundance ions. A linear improvement in ion intensity was observed with the number of trapping/accumulation events using the SLIM trap, illustrating its potential for enhancing the sensitivity of low abundance or targeted species.

Advancing the high throughput identification of liver fibrosis protein signatures using multiplexed ion mobility spectrometry.

Rapid diagnosis of disease states using less invasive, safer, and more clinically acceptable approaches than presently employed is a crucial direction for the field of medicine. While MS-based proteomics approaches have attempted to meet these objectives, challenges such as the enormous dynamic range of protein concentrations in clinically relevant biofluid samples coupled with the need to address human biodiversity have slowed their employment. Herein, we report on the use of a new instrumental platform that addresses these challenges by coupling technical advances in rapid gas phase multiplexed ion mobility spectrometry separations with liquid chromatography and MS to dramatically increase measurement sensitivity and throughput, further enabling future high throughput MS-based clinical applications. An initial application of the liquid chromatography--ion mobility spectrometry-MS platform analyzing blood serum samples from 60 postliver transplant patients with recurrent fibrosis progression and 60 nontransplant patients illustrates its potential utility for disease characterization.