High-Flow LC/MS Analysis of Immunosuppressant Drugs in Human Blood Using FAIMS Technology

Importance of the Topic

Monitoring immunosuppressant drugs in blood post-transplantation is critical to balance therapeutic efficacy and avoid toxicity. LC/MS methods face challenges from complex blood matrices that cause ion suppression and instrument contamination, reducing throughput. Incorporating FAIMS technology at high flow rates addresses these issues by enhancing selectivity and instrument uptime.

Objectives and Study Overview

This study evaluates FAIMS Pro coupled with high-flow LC/MS for quantifying four key immunosuppressants—sirolimus, tacrolimus, everolimus, and cyclosporin A—in human blood. The goal is to compare performance, robustness, and throughput against a conventional LC/MS setup without FAIMS.

Methodology and Instrumentation Used

• Sample preparation: Protein precipitation with zinc sulfate, spiking of analytes and ascomycin internal standard, centrifugation, supernatant analysis.
• LC conditions: Vanquish UHPLC Horizon at 0.8 mL/min, 3-minute run time, 15 µL injection volume.
• MS parameters: Orbitrap Exploris 240 in full scan (m/z 700–1400) at 60,000 resolution; HESI source optimized for auxiliary, sheath, and sweep gases; transfer tube and vaporizer temperatures tuned.
• FAIMS Pro: Compensation voltages empirically optimized inline for each analyte to filter contaminants and improve signal-to-noise.
• Data analysis: Thermo Scientific TraceFinder 5.1.SP2, assessing raw peak areas, response ratios, linearity, and precision over 5 days (1800 injections).

Main Results and Discussion

• Robustness: Without FAIMS, analyte peak areas declined >20% after three days; with FAIMS, signal remained stable over five days despite slightly lower intensity due to electrode transmission losses.
• Precision: %RSD of peak areas improved from >17–21% without FAIMS to <5% with FAIMS across all analytes.
• Linearity: Calibration curves (R² ≥ 0.99) remained consistent for both setups over five days, supporting reliable quantification.
• System stability: Fore-vacuum pressure remained constant with FAIMS, indicating reduced contamination and no need for mid-run maintenance.
• Response ratios: Internal standard correction produced stable analyte-to-IS ratios in both setups, highlighting the importance of internal standardization.

Benefits and Practical Applications

• Enhanced instrument uptime: Continuous operation for at least five days (360 injections/day) without cleaning stops.
• Improved data quality: Higher signal-to-noise and precision facilitate accurate therapeutic drug monitoring.
• Increased throughput: High-flow conditions and FAIMS integration reduce cycle time and maintenance downtime for clinical labs.

Future Trends and Potential Applications

• Broader adoption of FAIMS in high-throughput clinical and industrial workflows.
• Extension to other challenging bioanalyses requiring robust matrix cleanup inline.
• Further optimization of FAIMS parameters for multiplexed assays.
• Integration with automated sample preparation for fully hands-off therapeutic drug monitoring.

Conclusion

FAIMS Pro integration with high-flow LC/MS significantly improves robustness, precision, and throughput for immunosuppressant drug analysis in blood. This approach minimizes instrument downtime and enhances data reliability, making it well suited for routine clinical and research applications.

References

No external references were provided in the original text.