High-Throughput and Highly Selective Quantitative Lipidomics with the Stellar Mass Spectrometer – A Novel Hybrid Nominal Mass Instrument

Significance of the Topic

Rapid and precise quantification of diverse lipid species is critical for understanding metabolic disorders, biomarker discovery, and clinical diagnostics. Traditional workflows often struggle with throughput, isomer resolution, and specificity, limiting their application in high-volume and translational studies.

Study Objectives and Overview

This work evaluates the novel Thermo Scientific Stellar mass spectrometer in a targeted lipidomics workflow. Key aims include:

• Expanding lipid coverage and annotation specificity compared to established SRM methods
• Increasing throughput using parallel reaction monitoring (PRM) on a hybrid nominal mass instrument
• Demonstrating enhanced sensitivity and selectivity for clinical and translational research

Methodology and Instrumentation

Sample Preparation and Standards

• NIST SRM 1950 human plasma spiked with UltimateSPLASH ONE internal standards
• Protein precipitation and lipid extraction using isopropanol

Chromatography and Mass Spectrometry

• HILIC separation on ACQUITY Premier BEH Amide column (100×2.1 mm, 1.7 μm) with Vanquish Duo UHPLC
• Targeted PRM acquisition on the Stellar mass spectrometer combining a quadrupole and a dual-cell linear ion trap

Data Processing Tools

• LipidCreator for automated transition generation
• Skyline-Daily and PRM Conductor for method optimization
• R for statistical analysis and visualization

Key Results and Discussion

• Lipid coverage increased by over 50%, from ~1,019 SRM precursors to more than 5,300 transitions across positive and negative modes
• Peak sampling density improved by at least 20%, yielding average data points per peak of 10 (positive) and 15 (negative)
• Most lipid species now feature multiple transitions (average >3), enhancing confidence in isomer differentiation

These enhancements translated into more robust identification and quantitation of glycerophospholipids, sphingomyelins, and other classes within a 12-minute runtime, supporting high-throughput workflows.

Benefits and Practical Applications

• Higher selectivity and sensitivity accelerate biomarker discovery in clinical cohorts
• Parallel acquisition reduces cycle times, increasing laboratory throughput
• Improved isomer resolution aids in understanding lipid-specific biological roles

Future Trends and Potential Applications

Avenues for further development include:

• Implementation of targeted MS3 workflows for enhanced structural characterization
• Integration with automated sample handling for large-cohort studies
• Extension to multiomic platforms combining lipidomics with proteomics and metabolomics

Conclusion

The Stellar mass spectrometer, coupled with LipidCreator and optimized PRM methods, delivers a high-throughput, highly selective lipidomics platform. This approach overcomes key limitations of traditional SRM assays, offering increased coverage, sensitivity, and isomer resolution for translational and clinical research.

References

1. Remes et al. BioRxiv preprint, 2024. https://doi.org/10.1101/2024.05.31.596848
2. Medina et al. Analytical Chemistry, 2023, PMID: 36716250
3. Adams et al. Journal of Proteome Research, 2020, PMID: 31984744
4. Peng et al. Nature Communications, 2020, PMID: 32345972