LC-MS/MS Analysis of Immunosuppressant Drugs in Whole Blood using the Xevo TQ Absolute with Capitainer B qDBS Devices and Volumetric Absorptive Microsampling for Clinical Research

Significance of the Topic

Accurate monitoring of immunosuppressant drugs such as cyclosporine, everolimus, sirolimus and tacrolimus is essential in transplant medicine and autoimmune disease management. Traditional venous blood collection poses logistical challenges, including patient travel, invasive procedures and the need for trained personnel. Volumetric absorptive microsampling offers a minimally invasive alternative, enabling remote or home sample collection, reducing patient burden and expanding clinical research capabilities.

Study Objectives and Overview

The primary goal was to develop and validate a single-run LC-MS/MS method for quantifying four key immunosuppressants in 10 µL whole blood collected via Capitainer B qDBS devices. Performance metrics included analytical sensitivity, linearity, precision and accuracy, assessed against established quality assurance criteria.

Methodology

Samples of 10 µL whole blood were obtained using Capitainer B qDBS volumetric devices. An in-house LC-MS/MS protocol employed protein precipitation and solvent extraction containing stable isotopic internal standards. Chromatographic separation was achieved on a C18 column with a water/methanol/ammonium acetate gradient, yielding run times under two minutes. Quantification was based on multiple reaction monitoring in positive electrospray mode. Calibration and control samples from Waters MassTrak immunosuppressant sets and external quality assurance materials were included to evaluate method performance.

Instrumentation

• Waters ACQUITY UPLC I-Class System
• Waters Xevo TQ Absolute Triple Quadrupole Mass Spectrometer
• Capitainer B qDBS Volumetric Absorptive Microsampling Devices
• Waters C18 HSS SB Analytical Column

Main Results and Discussion

The method demonstrated limits of quantification of 1 ng/mL for everolimus, sirolimus and tacrolimus, and 25 ng/mL for cyclosporine with signal-to-noise ratios exceeding 10. Linearity was confirmed over 1–30 ng/mL (r2 > 0.995) for everolimus, sirolimus and tacrolimus, and 25–1500 ng/mL (r2 > 0.995) for cyclosporine. Intra- and inter-run precision (n=25) remained below 15% CV across low, mid and high concentration levels. External quality assurance samples met acceptance criteria with mean bias under 15%. These findings indicate robust assay performance comparable to standard venous methods.

Benefits and Practical Applications

Volumetric absorptive microsampling simplifies patient participation by eliminating venous access and travel to clinical sites. Key advantages include:

• Reduced invasiveness and improved patient comfort
• Enabling remote or at-home sample collection
• Minimized sample volume requirements (10 µL)
• High throughput with rapid LC-MS/MS analysis (<2 min per run)

This approach supports longitudinal clinical trials, therapeutic drug monitoring in decentralized settings and quality of life improvements for chronic patients.

Future Trends and Opportunities

Advances in microsampling technologies and portable mass spectrometers may further decentralize pharmacokinetic studies and therapeutic drug monitoring. Integration with telemedicine platforms and automated sample tracking could streamline remote patient management. Expanding the analyte panel, incorporating dried blood spot–based biomarker assays, and validating in diverse patient populations represent promising directions.

Conclusion

The validated LC-MS/MS method using Capitainer B volumetric devices achieves reliable quantification of cyclosporine, everolimus, sirolimus and tacrolimus in 10 µL whole blood. It meets stringent sensitivity, linearity and precision criteria while offering significant practical benefits for clinical research and decentralized therapeutic monitoring.

References

• Harrsch P. LC-MS/MS Analysis of Immunosuppressant Drugs in Whole Blood using the Xevo TQ Absolute with Capitainer B qDBS Devices and Volumetric Absorptive Microsampling for Clinical Research. Waters Corporation.