Direct quantitation of five immunosuppressant drugs in volume-controlled dried whole blood spots by a fully automated Transcend DSX-1 system

Importance of the topic

Dried blood spot (DBS) analysis for therapeutic drug monitoring of immunosuppressants addresses critical needs in transplant patient care by ensuring drug concentrations remain within therapeutic windows. Traditional immunoassays have given way to mass spectrometry techniques offering higher sensitivity and selectivity. Volume-controlled DBS sampling minimizes patient discomfort and simplifies logistics while maintaining analyte stability during storage and transport.

Goals and overview

This study demonstrates a fully automated end-to-end workflow for direct quantitation of five key immunosuppressant drugs (cyclosporin A, tacrolimus, sirolimus, everolimus, mycophenolic acid) from precise 10 µL dried whole blood spots. The Transcend DSX-1 system integrates flow-through desorption (FTD™) and 2-D liquid chromatography with TurboFlow™ technology, coupled to a TSQ Altis™ Plus triple quadrupole mass spectrometer, to achieve rapid and reliable analysis.

Methodology and instrumentation

Standard solutions spiked into K2-EDTA stabilized human whole blood covered calibration ranges from sub-ng/mL to low µg/mL levels. DBS samples were collected on HemaXis™ DB10 cards, air-dried, then processed by the Transcend DSX-1 dried spot module using an 8 mm clamp head. Internal standards were applied in-line, and flow-through desorption extracted analytes without manual punching. The extract underwent online TurboFlow clean-up before analyte focusing on a Hypersil GOLD™ C8 column and separation by a 2-step gradient. SRM detection on the TSQ Altis Plus MS with heated electrospray ionization achieved selective quantitation.

Used instrumentation

• Thermo Scientific Transcend DSX-1 system with dried spot module
• HemaXis DB10 volume-controlled blood collector
• Spark Holland flow-through desorption (FTD™) and Intelligent Vision Camera (IVC™)
• TurboFlow™ Cyclone-P column
• Thermo Scientific Hypersil GOLD C8 analytical column
• Thermo Scientific TSQ Altis Plus triple quadrupole mass spectrometer
• Aria MX and TraceFinder software suites

Main results and discussion

Total run time per spot was five minutes. Limits of detection and quantification ranged from 0.31 ng/mL to 6.25 ng/mL (LOD) and 1.25 ng/mL to 12.5 ng/mL (LOQ) across the drug panel. All calibration curves exhibited excellent linearity (R² > 0.99). Intra- and inter-spot precision (%RSD of internal standards) remained below 9%, and average bias at LOQ levels fell within ±15%. The automated workflow eliminated manual punching and extensive sample prep, reducing hands-on time and potential for error.

Benefits and practical applications

• Rapid, high-throughput quantitation suitable for clinical TDM laboratories
• Enhanced chain of custody with pre- and post-extraction imaging
• Reduced sample preparation labor and consumables
• Applicable to decentralized or remote sampling scenarios

Future trends and possibilities

Expanding the analyte panel for broader immunosuppressant and small molecule coverage could further support personalized medicine. Integration with decentralized point-of-care devices and digital data platforms may enable near-real-time dosing adjustments. Advances in microfluidics, AI-driven data analysis, and miniaturized MS sources could streamline workflows and reduce sample volumes even further.

Conclusion

The fully automated Transcend DSX-1 workflow provides a robust, reproducible, and high-throughput solution for quantifying multiple immunosuppressant drugs from volume-controlled DBS. This approach offers significant advantages in speed, accuracy, chain of custody, and operational efficiency, supporting improved patient management in transplant and other clinical settings.

References

1. Wu I, Guo J, Zhu Y, et al. Fully automated dried spot analysis for the rapid quantitation of tramadol and its metabolites in various matrices. Thermo Fisher Scientific Technical Note 000625, 2021.
2. Dollard SC, Dreon M, Hernandez-Alvarado N, et al. Sensitivity of dried blood spot testing for detection of congenital cytomegalovirus infection. JAMA Pediatr. 2021;175(3):e205441.
3. Guo J, Gibson RJ, Wu I, et al. Direct quantitation of phosphatidylethanol (PEth) in volume-controlled dried whole blood spots using the fully automated Transcend DSX-1 system. Thermo Fisher Scientific Technical Note 000301, 2022.