Cross-continental, multisite round robin REIMS study for the evaluation of REIMS fundamentals and technology

Importance of the Topic

Rapid evaporative ionization mass spectrometry (REIMS) enables real-time molecular profiling by analyzing aerosols produced during thermal ablation of biological samples. This in situ technique supports applications ranging from surgical margin assessment to food analysis and microbial identification on a quasi real-time basis. Establishing its reproducibility and robustness across multiple sites and instruments is critical for routine deployment in clinical and industrial settings.

Aims and Study Overview

• Evaluate repeatability and reproducibility of REIMS measurements across four institutions and six instruments.
• Assess instrument-to-instrument and site-to-site variability using standardized samples.
• Investigate key experimental parameters and fragmentation mechanisms to inform method standardization.

Methodology and Instrumentation

Six Xevo G2-XS mass spectrometers equipped with the iKnife REIMS interface were deployed at four locations: Maastricht University (M4I), Imperial College London (ICL), Queens University Kingston (Queens), and Waters Research Center Budapest (WRC). Samples included:

• Lockmass solution of leucin-enkephalin in isopropanol (0.05 ng/µL, 150 µL/min).
• NIST reference meat homogenate.
• Pork liver batches shipped from the UK and local meat (calf liver, chicken liver, chicken breast, turkey breast).

Data were acquired over multiple days by different users. Instrument parameters were harmonized via a checklist, including collision surface temperature, ion guide settings, and lockmass correction. Spectra underwent background subtraction and binning (0.1 Da) followed by multivariate analysis (PCA, LDA) and cross-validation for classification performance.

Main Results and Discussion

• Lockmass fragment ions showed consistent intensities across sites, indicating minimal fragmentation or dimer formation at the REIMS interface.
• First batch of pork liver samples revealed variability in fatty acid to phospholipid signal ratios between sites, likely due to pre-analytical differences.
• Second batch, after parameter adjustment, exhibited improved consistency, though minor site-specific differences persisted.
• Cross-site classification models achieved correct classification rates ranging from 64 % to 100 %, with WRC-based models demonstrating the highest cross-validation performance.
• Use of a common NIST reference provided a robust metric for instrument performance and data comparability.

Benefits and Practical Applications

The study confirms that REIMS can deliver reproducible and robust molecular profiles across continents and users. Key practical implications include:

• Facilitating real-time surgical margin assessment in oncology.
• Enhancing food authenticity and safety testing by rapid tissue classification.
• Supporting microbial and lipidomic analysis without extensive sample preparation.

Future Trends and Opportunities

• Development of standardized protocols and reference materials for global REIMS networks.
• Integration with real-time decision support and machine learning algorithms for automated classification.
• Miniaturized and portable REIMS devices for point-of-care and field applications.
• Expansion into new domains such as environmental monitoring and metabolomic profiling.

Conclusion

This cross-continental, multisite round robin study demonstrates that REIMS technology exhibits satisfactory repeatability, reproducibility, and robustness across different instruments and locations. While pre-analytical and site-specific factors can influence lipid profiles, harmonized instrument settings and use of standard references mitigate these effects, paving the way for broader clinical and industrial implementation.

References

• Balog J, Vaysse PM, Porta Siegel T, Kaufmann M, Amgheib A, Savage A, Varga V, Marton A, Pringle S, Rudan J, Heeren RMA, Takáts Z. Cross-continental, multisite round robin REIMS study for the evaluation of REIMS fundamentals and technology. Waters Corporation; 2019.