Combined PaperSpray and FAIMS technology for rapid quantification of immunosuppressants in whole blood for clinical research

Significance of the Topic

The quantified measurement of immunosuppressant drugs in whole blood is essential for maintaining therapeutic efficacy and minimizing toxicity in transplant and autoimmune patients. Traditional immunoassays can suffer from cross-reactivity and limited dynamic range, while chromatographic methods add time and solvent waste. A rapid, robust approach enhances clinical research workflows and patient care.

Objectives and Study Overview

This technical evaluation demonstrates the synergy between PaperSpray mass spectrometry and Field Asymmetric Ion Mobility Spectrometry (FAIMS) for the rapid quantification of cyclosporin A, tacrolimus, and everolimus in whole human blood. The study compares analytical performance with and without the FAIMS Pro interface to assess improvements in sensitivity, selectivity, and throughput.

Methodology

• Spiking whole human blood with calibration levels: cyclosporin A (10–1600 ng/mL), tacrolimus (0.5–80 ng/mL), everolimus (2.5–80 ng/mL) along with respective isotopically labeled internal standards.
• Spotting 10 µL onto single-use paper strips housed in VeriSpray sample plates and oven-drying at 45 °C for 30 minutes.
• Extracting analytes on-board with a 60% methanol/40% chloroform/0.1% sodium acetate solvent system, followed by PaperSpray ionization.
• Acquiring SRM data on a Thermo Scientific TSQ Altis triple quadrupole mass spectrometer, with optimized transitions and compensation voltages determined by FAIMS CV scans.
• Comparing runs with and without the Thermo Scientific FAIMS Pro interface to evaluate background reduction and LOQ enhancements.

Used Instrumentation

• Thermo Scientific VeriSpray PaperSpray ion source and plate loader
• Thermo Scientific FAIMS Pro interface
• Thermo Scientific TSQ Altis triple quadrupole mass spectrometer
• Thermo Scientific Xcalibur and TraceFinder software for data acquisition and integration

Main Results and Discussion

• FAIMS Pro integration sharply reduced matrix background signals (up to 100-fold), significantly boosting signal-to-noise ratios.
• Limits of quantitation improved from 10 to 0.5 ng/mL for tacrolimus and from 20 to 5 ng/mL for everolimus; cyclosporin A LOQ persisted at 25 ng/mL due to ion ratio criteria rather than S/N.
• Calibration curves showed excellent linearity, precision (<15% RSD), and accuracy (within ±20%) across therapeutic ranges.
• Representative chronograms revealed nearly complete removal of interferences at LOQ levels when FAIMS was applied, while direct PaperSpray without FAIMS exhibited higher baseline noise and poorer low-level accuracy.

Benefits and Practical Applications

• Minimal sample preparation and no chromatography enable analysis times under two minutes per sample.
• Automated plate handling allows unattended processing of up to 240 samples, supporting high-throughput clinical studies.
• Sensitivity and selectivity meet or exceed requirements for therapeutic drug monitoring in clinical research settings.

Future Trends and Potential Applications

• Extension of PaperSpray-FAIMS workflows to other therapeutic drug classes and biological matrices (urine, plasma).
• Integration into fully automated laboratory platforms for seamless, hands-free analyses.
• Advancements in FAIMS resolution and multiplexed ion separation to broaden analytical scope.
• Development of portable, point-of-care mass spectrometry systems for on-site therapeutic monitoring.

Conclusion

The combined use of VeriSpray PaperSpray ionization and FAIMS Pro interface offers a rapid, sensitive, and selective method for quantifying critical immunosuppressants in whole blood. By eliminating extensive sample cleanup and reducing background interferences, this approach streamlines clinical research workflows and holds promise for future diagnostic and point-of-care applications.

References

1. Walker K., Boeser C., Snyder R.A., Wijeratne N., Bhattacharyya D. Combined PaperSpray and FAIMS technology for rapid quantification of immunosuppressants in whole blood for clinical research. Technical Note 73307. Thermo Fisher Scientific; 2019.