lcmsIn 2013, we changed the paradigm for how our customers were able to approach their research with the introduction of the first Thermo Scientific Orbitrap Tribrid Mass Spectrometer (MS). The Orbitrap Tribrid architecture is truly unique with three mass analyzers; the Orbitrap mass analyzer enabling not only ultra-high resolution and mass accuracy, but also a sensitive and selective dual pressure linear ion trap that can perform parallel acquisition to maximize the experimental duty cycle, empowering our researchers to get more data with higher confidence and answer more questions.

Over the past six years, we’ve seen thousands of high-impact journal publications from our researchers redefining the experimental boundaries in multi-omics, translational research, and biopharmaceutical analysis to continue to accelerate scientific advancements. More than 1,500 publications covering more than 80 topics in the Omics space have been accepted since the release of the Orbitrap Fusion with the distribution shown below in the tree map.

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Everyday Performance Enhancements

The Thermo Scientific Orbitrap Eclipse Tribrid Mass Spectrometer extends the performance and experimental flexibility of the Orbitrap Tribrid architecture by introducing numerous key hardware and software advancements. Hardware improvements include a new quadrupole mass analyzer, improved vacuum manifold in the C-Trap/Orbitrap region, an extended high-pressure cell in the linear ion trap, and the option for Proton Transfer Charge Reduction source to perform gas-phase charge stripping. The new hardware features combine to support extended intelligence-driven instrument method templates that result in innovations that not only surpass previous instruments’ performance, but can again, change the paradigm of how research is accomplished. The data acquisition strategy with complex method templates ensures higher quality data for both experienced mass spectrometry-based researches as well as those just starting.

A great example of this is the enhanced quadrupole mass analyzer (Q1) increasing ion transmission over a wider mass range.  For existing bottom-up proteomic workflows, the new quadrupole mass analyzer maximizes ion transmission at narrow (0.4 m/z)   precursor isolation settings. The improved ion transmission significantly increases the number of unique peptides that can be identified from complex, biological samples resulting in greater characterization capabilities.

Advancing Tandem Mass Tag (TMT) Based Quantitation

Isobaric tagging strategies using Thermo Scientific™ Tandem Mass Tags™ Reagents (TMT™) are powerful tools for studying how proteins interact and function in biological systems. Up to 11 samples or more can be multiplexed in a single, high-resolution LC/MS experiment to enable state-of-the-art quantitative analysis of peptide and protein abundance.

Real-Time Search is a new feature which identifies and sequences peptides on the fly. For the first time, the Orbitrap Eclipse data acquisition strategy will intelligently determine which linear ion trap MS2 spectra will be carried out for a Synchronous Precursor Selection MS3 (SPS MS3).  In addition, to determine which precursors have matched a user-defined indexed database, the intelligent on-the-fly processing routine determines which specific set of product ions are isolated and dissociated increasing the accuracy and precision in peptide identifications quantitation and which ones won’t. The addition of Real-Time Search results not only significantly improves proteome coverage, and enhanced quantitative accuracy, but almost doubles sample throughput.

Proton Transfer Charge Reduction (PTCR) Option

PTCR is a new ion-ion reaction-based methodology enabling scientists to strip off protons through controlled, gas-phase reactions. This is exciting because it enables a sophisticated precursor/product ion purification processes making uninterpretable spectra, interpretable, especially for complex, large-scale, top down analyses.

The communities most excited about PTCR are the top-down proteomics, the biopharma community, and anyone else who’s analyzing large native complexes. The reason they’re interested is because now they can take these very complex samples that may contain numerous proteoforms (based on different post-translational modifications (PTMs), sequence variants, and/or truncation/clippings) differentiated by small mass differences and reduce their complexity. Performing the simplification step within the mass spectral experiment saves time, reduces sample perturbation, and increases researchers’ ability to achieve more complete and more confident sequence assignment or confirmation of proteins and protein complexes.

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Extended High Mass Rangen (HMRn) Option

The new Orbitrap Eclipse MS instrument design and additional functionality extends the MS and MSn ranges to m/z 8000. This improvement allows for top down MSn experiments over a wider mass range, than currently achievable. It is now possible to structurally analyze individual proteins and protein complexes in their native state. This will enable scientists to tear apart large complexes and native proteins in ways they were not possible before, leveraging the various dissociation techniques only offered on the Tribrid platform in a way that’s really exciting the biopharma community and others.

Professor Dame Carol Robins Research Group at the University of Oxford has pioneered using native mass spectrometry to study membrane protein assemblies to understand how the proteins that form large assemblies come together and how interactions with small molecule ligands can change the structure and confirmation of those assemblies. The biology of membrane proteins is really challenging to study because these assemblies are surrounded in their native environment by lipids and other molecules that enhance their stability. As soon as you remove them from this lipid bilayer they lose their stability and also the transient interactions with other molecules that you might wish to capture.

The Orbitrap Eclipse Tribrid MS allows researchers to visualize intact membrane protein-ligand assemblies but also to dissect them into their protein and ligand components. Using the MSn capability of the instrument, scientists can now define proteoforms that make up membrane protein complexes and provide chemical identity on the ligands that bind in a unified workflow.

Greater Sequence Coverage for Intact Protein Analysis

In 2017, Ultraviolet Photodissociation (UVPD) option was introduced  on the Orbitrap Fusion Lumos Tribrid Mass Spectrometer to enable complementary fragmentation for intact protein analysis and allowed greater sequence coverage, resulting in faster data analysis, as well as higher confidence to understand not only specific proteins, but also understand the differences or the appearance of proteoforms.

The Orbitrap Eclipse is also capable of combining UVPD and extended Electron Transfer Dissociation (ETD) options to increase the way users can characterize not only peptides or modified peptides, but also expand their capabilities for intact protein analysis in top down — making it easier for classic proteomic applications but also enabling more effective and efficient biotherapeutic characterization, as well as structural biology in terms of protein-protein cross linking.

Summary

With a continued commitment to deliver advanced performance capabilities, the Thermo Scientific Orbitrap Eclipse Tribrid mass spectrometer expands experimental methods; one system for maximum insights. Researchers will be able to acquire more data with higher confidence in less time and with less sample and surpass today’s research challenges.

Learn more at thermofisher.com/orbitrapeclipse

Publication: Active instrument engagement combined with a real-time database search for improved performance of sample multiplexing workflows