MSACL 2025 Abstract

Evaluation of an Ultra-High Throughput Method for the Analysis of Tacrolimus in Whole Blood using the SCIEX Echo MS+ System

Michael J. Y. Jarvis (1), Sarah Delaney (2), Leonard Chay (1), John Gibbons (1)
(1) SCIEX, Concord, Ontario, Canada, (2) Unity Health, Toronto, Ontario, Canada

Michael Jarvis (Presenter) SCIEX

Relevant Financial Disclosures (within past 24 months, reported on Jul 17, 2025) Other Potential Conflicts SCIEX / Employee Conflict mitigation NOT REQUIRED.

INTRODUCTION:
The SCIEX Echo MS+ system utilizes acoustic droplet ejection technology to provide ultra-high throughput, carryover-free analysis of samples from a variety of aqueous matrices. This technique is typically most attractive for assays that have large sample volumes, due to the speed (up to 3 Hz) of the sample analysis. In this study we have chosen to evaluate the applicability of the Echo MS+ system for the analysis of Tacrolimus. MS analysis is widely considered the gold standard for highly accurate, highly specific quantitation of immunosuppressant drugs. The attractiveness of this ultra-rapid technique for the analysis of immunosuppressant drugs such as Tacrolimus is not only its throughput, but also its ability to deliver rapid, same-day results.

METHODS:
The analysis of Tacrolimus was performed in pure solvent and in whole blood samples, after appropriate sample preparation. Whole blood calibrators and controls were obtained from Diagnotix (Appingedam, Netherlands). For the whole blood samples, calibrators, and controls, 100uL of sample was added to 200uL of concentrated ZnSO4 solution, the samples were vortex mixed, then allowed to incubate at room temperature for 5 minutes, after which the samples were again vortex mixed and finally centrifuged at 10,000 g. The clean supernatant was transferred to a 384-well microtiter plate. Sample injection (or “ejection”) using acoustic droplet ejection technology on the SCIEX EchoMS+ system uses sound energy to causes 2.5nL droplets to be ejected from the well plate for capture in the “open port” probe, which transfers the sample to the mass spectrometer source. In this study, we have evaluated sample ejections ranging from 1 droplet (2.5nL) up to 400 droplets (1uL) per sample. Regardless of the number of droplets ejected, the sample-to-sample injection time was always <5 seconds. MS/MS analysis was performed using electrospray ionization in positive mode on the SCIEX Triple Quad 6500+ system.

RESULTS:
A variety of parameters were evaluated to determine optimum method conditions, including carrier solvent composition, carrier solvent flow rate, carrier solvent modifiers, ion source conditions, and the number of droplets per injection. Our results demonstrate that although a higher percentage of methanol or acetonitrile in the carrier solvent provides enhanced ionization (raw signal, measured in counts-per-second) for Tacrolimus, there is an increase in the background that negatively impacts the S/N, therefore it is preferable to operate with predominantly aqueous carrier solvent. Increasing the ejection volume has the effect of increasing the signal, and hence the LOQ, however it also results in slightly wider peaks. Nevertheless, even with ejection volumes as large as 1000nL we were able to easily achieve sample-to-sample injection times of less than 5 seconds. A linear response was achieved over a concentration range from 1 ng/mL to 1000 ng/mL of Tacrolimus, with %CVs <15% and accuracy +/- 15% across the entire concentration range. Tacrolimus concentrations determined in whole blood samples on the Echo MS+ system were compared to measurements obtained by immunoassay on the same samples, and demonstrated a good correlation.

CONCLUSION:
In this study we have successfully demonstrated the applicability of the ultra-high throughput Echo MS+ system to the measurement of Tacrolimus in whole blood samples. The work presented here should be readily extensible to the measurement of additional immunosuppressant drugs such as Cyclosporine A, Everolimus, and Sirolimus in whole blood. The method presented here demonstrated a linear response across a concentration range from 1 ng/mL to 1000 ng/mL of tacrolimus, with acceptable %CVs and Accuracy% across the measured concentration range. A direct comparison of Tacrolimus measurements in whole blood samples using the method on the Echo MS+ system demonstrated a good correlation with measurements obtained via immunoassay on the same samples.