Revolutionizing translational research: large-scale targeted PRM proteomics assays enabled by the Stellar mass spectrometer

Importance of the topic

Recent advances in targeted proteomics enable high-throughput biomarker verification in clinical research at unprecedented scale.

Objectives and Study Overview

This work demonstrates a streamlined workflow using the Stellar MS, Skyline, and PRM Conductor to develop and validate large-scale PRM assays for hundreds of plasma proteins within 3–4 days.

Methodology and Instrumentation

Sample preparation: Biognosys PQ500 heavy reference peptides spiked into digested human plasma at 300 ng/µL, with an 11-point serial dilution series to assess LOD, LOQ, and linearity.
Liquid chromatography: Thermo Scientific Vanquish Neo UHPLC with EASY-Spray ES906A column, trap-and-elute injection, 45 °C, using 60 SPD (24 min) and 100 SPD (14 min) gradients.
Mass spectrometry: Thermo Scientific Stellar MS featuring high-resolution accurate mass linear ion traps, dynamic maximum injection times per target concurrency, Adaptive RT real-time chromatogram alignment, and PRM Conductor external tool within Skyline for automated method creation.

Main Results and Discussion

The transition list workflow generated scheduled PRM methods comprising 1 622 precursors and ~13 800 transitions for both 60 SPD and 100 SPD methods.
Performance metrics: 94 % of peptides achieved CV < 20 % (n = 10); LOQ for most peptides < 50 amol, LOD < 50 amol; the 60 SPD method yielded slightly lower LOD and LOQ than 100 SPD.
Automated filtering based on signal-to-noise, peak area, retention time, and transition consistency combined with adaptive scheduling ensured robust quantitation in plasma.

Benefits and Practical Applications

This platform offers enhanced sensitivity, specificity, and quantitative accuracy for highly multiplexed targeted proteomics, supporting large-scale biomarker verification and translational research in complex biological matrices.

Future Trends and Potential Applications

Integration of AI-driven method optimization, expanded peptide panels, multi-omics workflows, and clinical laboratory implementation is expected to further accelerate targeted proteomics applications.

Conclusion

The combination of Stellar MS, Skyline, and PRM Conductor delivers a rapid, scalable, and precise solution for large-scale PRM assays, facilitating translational biomarker studies with high sensitivity and reproducibility.

References

[1] PanoramaWeb. PRM Conductor resource. https://panoramaweb.org/prm_conductor.url
[2] Remes PM, Jacob CC, et al. bioRxiv Preprint 2024. doi:10.1101/2024.05.31.596848