Orbitrap Ascend Structural Biology Tribrid mass spectrometer - Product specifications

Importance of the Topic

Mass spectrometry has become a cornerstone technique in structural biology and proteomics, enabling detailed characterization of intact proteins, post-translational modifications, and native complexes. High sensitivity, rapid scan rates, broad mass range, and versatile fragmentation methods are essential to meet the growing demands of modern biopharmaceutical research, quality control, and fundamental studies of biomolecular assemblies.

Objectives and Study Overview

This application note presents the Thermo Scientific Orbitrap Ascend Structural Biology Tribrid mass spectrometer and highlights its ability to scale up native structure analysis. The goal is to demonstrate the instrument’s performance in terms of sensitivity, throughput, mass accuracy, resolution, fragmentation versatility, and automation features that support complex workflows from native mass analysis to quantitative proteomics.

Methodology and Instrumentation

The Orbitrap Ascend integrates several novel hardware components and software functions to achieve exceptional performance:

• Active Ion Management (AIM+) with QR5 segmented quadrupole to maximize ion transmission and selectivity across m/z 50–8000.
• Dual Ion Routing Multipoles (IRMs) for flexible storage, dissociation (HCD), and transfer to the Orbitrap or linear ion trap.
• Ultra-high-field Orbitrap mass analyzer delivering up to 480,000 FWHM resolution at m/z 200 and sub-ppm mass accuracy with internal calibration.
• Dual-pressure linear ion trap for CID, ETD/EThcD/ETciD, PTCR, and optional UVPD, supporting MSn up to n=10 with acquisition rates up to 50 Hz.
• Extended mass range native MS option up to m/z 16,000 for intact protein complexes.
• Thermo Scientific Ion Max NG and Auto-Ready ion sources for hands-free calibration and robust electrospray or APCI/APPI operations.
• Optional hardware modules including EASY-ETD, PTCR, UVPD, FAIMS Pro Duo, and NanoSpray/EASY-Spray interfaces.
• Comprehensive software ecosystem: Xcalibur control, Real-Time Search, Dynamic Maximum Injection Time, Advanced Peak Determination, TurboTMT, AcquireX, Proteome Discoverer, Compound Discoverer, and BioPharma Finder.

Main Results and Discussion

Performance tests using standard calibration solutions demonstrate:

• MS/MS acquisition rates of up to 45 Hz in the Orbitrap and 50 Hz in the linear ion trap with short maximum fill times.
• Sensitivity for 100 fg reserpine with S/N >200:1 in ESI MS/MS mode at 500 µL/min.
• Mass accuracy better than 1 ppm RMS over 24 hours with internal calibration.
• Dynamic range exceeding 5000 within a single high-resolution spectrum.
• Efficient ETD, PTCR, and UVPD fragmentations with high product-ion yields (≥15–25%).
• Native MS analyses capturing both precursor and product ions up to m/z 16,000, facilitating comprehensive characterization of protein assemblies.

These results confirm the system’s ability to handle both routine high-throughput proteomic experiments (e.g., TMT quantification with TurboTMT and Real-Time Search) and challenging native mass or cross-linking workflows without sacrificing speed or data quality.

Benefits and Practical Applications

The Orbitrap Ascend Structural Biology Tribrid offers:

• Unprecedented experimental throughput and flexibility across a wide range of applications from top-down proteomics to native complex analysis.
• Automated calibration and optimized method templates for consistent, unattended operation.
• Orthogonal fragmentation methods (CID, HCD, ETD, PTCR, UVPD) to maximize sequence coverage and structural insight.
• Real-Time Search and dynamic acquisition strategies to increase identification rates and quantitative accuracy in isobaric labeling experiments.
• Seamless integration with advanced data processing platforms for small-molecule profiling, targeted quantitation (SureQuant), and biotherapeutic characterization.

Future Trends and Potential Applications

Looking ahead, continued development in several areas will further enhance structural mass spectrometry capabilities:

• Integration of AI-driven acquisition algorithms for intelligent real-time decision making and adaptive workflows.
• Expansion of ion mobility separations (e.g., FAIMS or TIMS) for improved isomer separation and conformational analyses.
• Higher repetition-rate UVPD and improved photochemistry options for more comprehensive fragment coverage.
• Enhanced front-end microfluidics and automated sample handling to support single-cell and low-input proteomics.
• Deeper coupling with cloud-based data analytics and collaborative platforms for large-scale atlas-style projects.

Conclusion

The Thermo Scientific Orbitrap Ascend Structural Biology Tribrid mass spectrometer represents a significant advancement in mass spectrometry for structural biology and proteomics. Its combination of sensitivity, speed, resolution, and fragmentation versatility, paired with robust automation and integrated software, equips laboratories to tackle both current and future challenges in biomolecular characterization.

Used Instrumentation

• Orbitrap Ascend Structural Biology Tribrid mass spectrometer
• Ion Max NG electrospray and Auto-Ready calibration ion sources
• EASY-ETD, PTCR, and optional UVPD modules
• FAIMS Pro Duo interface
• Xcalibur and specialized data acquisition/processing software packages