Confident quantitation for every user: immunosuppressants in human blood by liquid chromatography-tandem mass spectrometry for clinical research

Význam tématu

Therapeutic monitoring of immunosuppressive drugs is essential to prevent graft rejection and drug toxicity in organ-transplanted patients. Liquid chromatography-tandem mass spectrometry offers superior selectivity and specificity compared to traditional immunoassays, enabling accurate quantification of multiple immunosuppressants in a single run.

Cíle a přehled studie

This study aimed to develop and validate a robust LC-MS/MS method for simultaneous quantification of four immunosuppressants—tacrolimus, sirolimus, everolimus, and cyclosporine A—in human blood using a Thermo Scientific TSQ Quantis triple quadrupole mass spectrometer coupled to a Transcend LX-2 UHPLC system.

Použitá metodika a instrumentace

Sample preparation:

• Offline protein precipitation of 100 µL blood with 0.1 M ZnSO4 and methanol containing isotopically labeled internal standards.
• Centrifugation and transfer of supernatant to analysis plate.

Chromatography:

• Thermo Scientific Transcend LX-2 dual-channel UHPLC.
• Hypersil GOLD column 50×2.1 mm, 1.9 µm, at 60 °C.
• Gradient elution with water and methanol (10 mM ammonium formate, 0.1% formic acid) and organic mixture of acetonitrile/acetone/2-propanol.
• Three-and-a-half minute runtime.

Mass spectrometry:

• Thermo Scientific TSQ Quantis triple quadrupole with heated electrospray ionization in positive mode.
• Selected-reaction monitoring with two transitions per analyte.
• Calibration with 1/x weighted linear interpolation.

Hlavní výsledky a diskuse

The method showed excellent linearity (R2 > 0.999) across calibration ranges, negligible carryover, accuracy bias within ±6.3%, and intra- and inter-assay precision with CVs below 7.3%. Representative chromatograms and calibration curves confirmed sensitivity at low nanogram-per-milliliter levels.

Přínosy a praktické využití metody

• Simple offline sample preparation and rapid analysis increase laboratory throughput.
• Simultaneous quantification of four key immunosuppressants in a single assay run.
• High sensitivity and precision support reliable therapeutic drug monitoring in clinical research.

Budoucí trendy a možnosti využití

Future developments may include integration with automated sample handling, expansion to additional drug panels, miniaturization for finger-prick blood samples, and real-time data processing with advanced software for individualized dosing algorithms.

Závěr

The presented LC-MS/MS method on the Transcend LX-2 and TSQ Quantis platform provides a rapid, sensitive, and precise approach for comprehensive immunosuppressant quantification in human blood, meeting clinical research requirements.

Reference

De Nardi C, Brambilla P, Brambilla M, Fania C. Confident quantitation for every user: immunosuppressants in human blood by liquid chromatography-tandem mass spectrometry for clinical research. Thermo Fisher Scientific Technical Note 65439; 2019.