Perspectives from ASMS 69: Part I

One of the products exhibited at ASMS 69 is the SELECT SERIES MRT by Waters, which includes Multiple Reflecting Technology to fold a 46-meter ion path into a compact vacuum chamber.

Due to COVID-19, ASMS 2021 was held as a virtual/in-person hybrid meeting. ASMS 69 needed a virtual component since travel to the U.S. was still restricted. Depending on country of origin, those outside the U.S. could face a multi-week quarantine upon returning home. New Zealand, for example, was set to impose a two-week quarantine for potential returning residents.

COVID-19 caused delays in many development programs due to shutdown of R&D facilities. I was interested in the impact on product and applications development—the loss was much less than I’d expected. For example, the staff at Bruker apparently used the shutdown to catch up on writing—they were listed as authors on 48 posters. My sampling shows the quality is very good.

Some statistics from ASMS:

* In-person= 2,784; Remote= 2,245

Notes from the Exhibition

A total of 112 firms exhibited at ASMS 69. However, most of the products that made the floor impressed me as extensions rather than new platforms. Ion mobility was an attention grabber, with instruments and also as a stage in the MS ion train.

Folded ToF Mass Analyzers

The folded ToF (time-of-flight) mass analyzers showed the most novelty in MS instrumentation. Multiple Reflecting Technology (MRT) from Waters provides a 46-meter ion path in a suitcase-size vacuum chamber. Designed for MS imaging, the SELECT SERIES MRT provides high mass resolution (~200,000) and ppb mass accuracy. The ability to use the isotopic fine structure to discriminate between isobaric analytes improves image sharpness. Ambiguous isobars are largely eliminated. Imaging is the first target application segment, but soon the MRT technology will be fitted with a ESI ion source for direct injection, or for uHPLC-uHRMS. No doubt the front end will be automated since Waters acquired Andrew Alliance. 

I asked about possible infringement with LECO’s folded W technology, which was introduced about five years ago for GCxGC. I was told that LECO is a licensee for GC x GC and Waters is licensed for imaging and ESI.

Bending the Tower

TOF mass spectrometers are usually identified by the characteristic long or tall drift tubes. Bruker’s three-story Max is an extreme case in point. Reflectrons cut the length in half, sometimes more with multiple passes. But, JEOL introduced a new TOF flight path where the ions traverse a horizontal figure 8 path that facilitates stuffing a high resolution TOF analyzer in a box about the size of a desktop personal computer of 1980 vintage. JEOL’s design uses multi-turn technology to bend the analytes’ flight path. The JMS-MT3010HRGA ("INFITOF") provides high mass resolving power (~30,000 FWHH) with a m/z range of 1 to 1000. It can detect H⁺, and resolve ions such as H₂⁺, D⁺, N₂⁺ and CO⁺. Target applications includemeasurement of high-purity gases in labs developing semiconductors, catalysts andhigh-end tech materials, petrochemical production (as in methane leaks) and refining plus monitoring exhaust gases.

Agilent replaced the 6545 MS with the 6560C Ion Mobility LC/QTOF, which is managed by a new release of VWorks Software. A morning lecture discussed its use in comprehensive and heart-cut LCxLC when coupled to Agilent’s 1290 series HPLC. Version 14 of the VWorks Software controls instrument and data functions plus report preparation. For HOS and PTM studies, the 6560C can be equipped with an ExD ion source from e-MSion (below) for electron capture dissociation (ECD).  ECD’s low energy ionization helps reduce post ionization isomerization of glycans and disulfides. The ExD induces side-chain fragmentation that is useful in differentiating isobaric amino acids, which can be important in structure/activity relationships.

Agilent’s HRdm is another neat feature of the 6560C that support multiplexing that magically increases the ion mobility resolution from 50 to 250. The HRdm uses a library of thousands of computational models to simultaneously probe for deconvolution and demultiplexing solutions. The output is a limited set of in-class isomers such as oligosaccharide isomers.

Additional MS Systems

• JEOL’s JMS-TQ4000 Triple-Quad mass spectrometer is designed for quantitative analysis. It boasts the highest MRM switching speed of 1000/second. This enables quantitation of many ions in a single run. To further improve throughput, the short collision cell accumulates ions that are pulsed into the mass analysis quadrupole. Pulsing is coordinated with the signal acquisition for optimum signal/noise. The fast quadrupole is compatible with very fast GC using short, narrow bore capillaries and rapid temperature programing.
• Sciex introduced the ZenoTOF as a revolution in MS, since it is capable of multiple fragmentation modes coupled with ion optics that pass more than 90% of the ions to the TOF mass analyzer. Plus, the duty cycle is increased to over 90%, which improves detection sensitivity by 4 to 20x. The key advance is the Zenotrap, which gathers and then ejects packages of ions into the mass analyzer. The improved detection sensitivity can be used to take smaller samples or dilute away matrix effects, which leads to more useful platform methods.

Portable Mass Spectrometers

In analytical instruments, maturity of a technology is often signaled by introduction of smaller portable instruments. The ASMS-69 technical program led off with a session focused on Portable LC-MS.

For example, John Crutchfield, iChrom Solutions, used a portable SPE-LC-MS system developed by Microsaic Systems to analyze water and soil samples for perfluorinated alkyl substances (PFAS). The assay worked quite well with detection limits for various analytes of about 3 ppt and calibration curves spanning 10 to 500ppt with triple 9 R² values.

BaySpec, Inc. introduced two linear ion trap mass spectrometers. While the mass analyzer is light and small, it requires a vacuum pump and power supply.  Ionization modes include electron impact (EI), thermal desorption (TD), electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI), and any other ambient ionization techniques such as DART and DESI.

ThermoFisher Scientific introduced a three member series of TSQ Plus Quadrupole mass spectrometers. Each features a bent ion beam guide to block uncharged analytes. In addition to the Delta Q (below), Thermo Scientific also introduced the Orbitrap IQ-X Tribrid mass spectrometer, The FAIMS Pro Duo interface andthe Vanquish Neo UHPLC System.

Thermo’s OrbitrapIQ-X Tribrid mass spectrometer is optimized for on-the-fly identification of small molecules such as drug metabolites. The key advance is that the Tribrid integrates Thermo’s AcquireX automated data acquisition capability which provides real time identification of peaks based upon MS/MS library search and matching. The IQ-X platform is compatible with several of Thermo’s ion sources such as Ultraviolet Photodissociation (UVPD) and the FAIMS Pro Duo interface.

Imaging with TOF-MS

MassTech Inc., developed atmospheric pressure MALDI inlets for imaging, particularly of biological specimens. At ASMS 69, attention focused on the MT Explorer 50, which is a compact MS designed for at-site chemical identification using MT’s ESI/APCIion source—or, the Explorer can use the ESI/DART/APCI/MALDI ion source. MassTech published a detailed method for “Mass spectral imaging using AP-MALDI ionization source coupled with Sciex QTOF 6600 analyzer.” The poster provides a step-by-step detail from source installation on the MS to sample preparation, including spraying the MALDI matrix, selection of target area and image preparation.

Add-on Modules

Excellims Corporation is a recognized vendor of stand-alone ion mobility spectrometers (IMS) plus IMS add-ons for mass spectrometers. The Excellims MA3100 IMS add on brings IMS to orbitraps. Options include liquid drift gas modifiers plus additional inlets for gas or solid samples. The add-on for MS is commonly an electrospray interface.

CovalX extend the Pearl high-mass detection module for MALDI-TOF analyzers with introduction of the HM4. This detector uses dynode technology to characterize chemical and biochemical complexes up to 2 MDa in size. Plus, the HM4 detects anions, as well as cations. The HM4 is designed to help characterize large protein complexes including post translational modifications, aggregates and mapping of disulfides and epitopes. Like the prior Pearls, the HM4 can be added to MALDI mass analyzers from Sciex, Bruker and Shimadzu. CovalX also provides MALDI-TOF characterization services using their instruments on a fee basis.

Thermo’s FAIMS Pro Duo MS interface provides differential ion mobility separation to further resolve overlapping peaks in LC-MS, or CE-MS. FAIMS, (an acronym for Field Asymmetric Ion Mobility Spectrometry,) is an interface to a mass analyzer designed to differentiate isobaric ions while passing through high and low strength electric field. It is most useful in assays of polysaccharides and lipids. The FAIMS pro Duo is designed to fit on Thermo Scientific’s Orbitrap Exploris; Tribrid series; TSQ Quantis; Altis; and TSQ Plus mass spectrometers. The PRO Duo interface is easy to install without breaking vacuum.

Ion Sources

Since helium needs to be conserved, Ionsense introduced their JumpShot Technology, which replaces the continuous flow of helium sweeping their DART-SVP with pulses of helium. This reduces He emission by up to 95%. It also improves the signal-to-noise and peak shape, while tripling throughput. Win, win, win.

e-MSion, Inc. featured the ExD ion source, which provides both electron induced and electron transfer dissociation (EID and ETD). Both are soft ionization techniques that generally give larger ions and thus more complete coverage than other dissociation modes such as collision-induced dissociation (CID). Plus, ExD is lower energy, and structural details such as the location of post translational modification (PTM) are preserved. ExD Cells are available for the Agilent LC-Q-ToF, Thermo’s Q Extractive Orbitrap MS series and the SELECT SERIES Cyclic IMS from Waters. Permanent magnets distribute the electrons around the detection cell.

CMP Scientific makes a sheath flow nanospray ion source (the EMASS-II CE-MS) designed for ‘omics applications, such as characterization of charge variants of biopolymers.

SESI (Secondary Ion Electrospray Ionization) is designed for characterization of vapors and nano aerosols. It is promoted by Fossilion Tech of Madrid, Spain. Vapors are heated and mixed with protonated water clusters produced by a nano electro spray and then transferred on-line to the mass spec such as an LTQ, or Extractive or Q Extractive Orbitrap from Themo Scientific.  In the U.S., Fossiliontech is represented by ESI Source Solutions.

Sample Prep and Pre-analytics

Until recently, digital microfluidics (DMF) has been limited to qualitative analysis in proteomics. A report from Jarrod Sandow of IonOpticks used DMF to evaluate  detergents of differentially expressed proteins in low cell number proteomes. He found that 3-dodecyloxypropyl-1-ß-D-maltopyrandoside was compatible with on-chip drop movement and sample preparation for quantitative assays of small cell count samples using LC-MS with a Q Extractive MS (ThermoFisher).

908 Devices introduced the ZipChip Oligos kit for qualitative and quantitative oligonucleotide analysis using CE-MS (capillary electrophoresis-MS). The kit provides “dilute and shoot” simplicity for oligos with various modifications. The MS provides accurate mass identification.

Speaking of oligos, Bruker’s new OligoQuest software in the GLP-ready BioPharma Compass suite offers enhanced RNA and oligonucleotide characterization. OligoQuest leverages high isotopic fidelity from maXis II and timsTOF Pro to characterize nucleic acid macromolecules, such as single-guide RNA as well as their impurities. Furthermore, fast PASEF acquisition maps complex mixtures derived from digested mRNA samples. Co-developed with RiboDynamics, LLC, OligoQuest offers algorithms and workflows to annotate tandem MS data from oligonucleotide sequences >100 nucleotides.

The trend to smaller samples inevitably leads to the need for new technology for solid phase extraction. Affinisep expanded its AttractSPE Disks for micro scale solid phase extraction. The kits use several formats for biomolecular applications, such as spin tubes, StageTips, 96 well plates etc. These employ discs that have dense packing of selective sorbent particles. The thin discs reduce elution volumes but are robust enough for centrifugation. Sorbent phases include C18, RPS, SDB, SCX and SAX. AttractSPE Disks BioMol is recommended for peptide purification including desalting of samples. The discs are an alternative to Empore SPE membranes. The latter are now marketed by CDS Analytical.

Interference from matrix components in biological samples is a common frustration. The Biotech Support Group introduced AlbuVoid and AlbSorb for albumin removal. Similarly, Cleanascite removes lipids, and HemogloBind removes hemoglobin, greatly simplifying the MS spectra. The firm works with other vendors for clinical assays for detection of biomarkers from specimens from COVID-19 and cancer patients.

Additional sample prep introductions include:

• Covaris specializes in Adaptive Focused Acoustics (AFA) to aid in preanalytical sample preparation. Using high-frequency acoustics is consistent with a tight focus of the acoustic energy with lower heating of the rest of the sample. This can be important for preserving the higher order structure of biomolecules and tissue specimens. The largest use of AFA energetics is shearing of DNA and RNA for next generation sequencing plus extraction of nucleic acids and proteins from FFPE samples. AFA is also used in cyro-pulverization, since it is quick and easier than most other sample grinding techniques.       
• Evosep introduced the EVOSEP ONE, which is an automated sample prep station for bioanalysis, including clinical diagnostics. The design goal is to be ten times faster and 100 times more robust. Most of the speed comes from UHPLC column technology (small diameter particles in short, nanobore columns) coupled to a fast MS. For robustness, Evosep is working with subject matter experts to improve workflows. For example, in the disease discovery stage, Evosep is working with Angela McArdale of Cedars-Sinai to develop a robust workflow using data independent acquisition (DIA) for high throughput for detection of marker proteins for COVID-19 infection and progression. At the other extreme, the EVOSEP ONE can be programed for rapid targeted assays with throughput up to 300 samples/day. At ASMS 69, Evosep and Sciex announced a comarketing agreement where Evosep focuses on the preanalytical stage and Sciex provides the MS, including software.
• Even for portable iMS instruments there is the issue of the last few inches between the mass analyzer and the sample specimen source. The analyte may be on a building surface, or the skin of an animal. Trace Matters Scientific (Somerville, MA) bridges this gap with a flexible low loss ion guide tube, called the SPion, with an orifice probe that gathers and transports ions down the center and into the mass analyzer. The device appears to be compatible with various ion generation technologies, such as field desorption, photo ionization and MALDI.
• PreOmics developed and introduced the BeatBox, which quickly and effortlessly homogenizes the samples in a single use 96-well plate format. Each well in the plate contains a single magnetic bead with a proprietary coating. A comparative study showed that the BeatBox gave higher protein extraction from 1 to 2 mg specimens. Reproducibility was also improved. Runtime is adjustable up to 10 minutes. After solubilization the plate is transferred to a rack that attracts the bead to the well side, enabling aspiration of the sample.
• Spraying the MALDI matrix on the sample is usually a manual operation, but this may be a significant contributing factor to the signal imprecision, even within a single sample plate. In operation, Shimadzu’s iMLayer AERO moves the plate at a uniform velocity in front of the fixed sprayer. This is repeated several times to provide a uniform coating of small matrix crystals. This improves the signal reproducibility as well as spatial resolution. The AERO has an automated rinsing cycle to prevent clogging. The humidity and temperature of the spray chamber are also controlled. The AERO is but one example of Shimadzu’s program to improve analytical reproducibility by reducing environmental variability and automating previously manual operations.

Single Cell Omics

Cell sorters have a 50+ year history, but Cellenionhas opened a new segment focused on single cell omics. Selection and manipulation of single cells is the first challenge. Cellomics uses a video-microscope focused on a fixed length of a capillary tip to reject trash, multiple cells and non-target cells, and select an aliquot containing a single cell of interest. Once identified, the tip is moved to the specific sample well in or on a multi-well plate. At ASMS 69, Cellenion introduced the proteoCHIP 12*16 for miniaturized sample preparation of single cell workflows.

Check back on Friday, Nov. 19 for Part II of Perspectives from ASMS 69, including inorganic applications, hybrid instruments, LC-MS columns and MS software.