Multiplex Immunosuppressant (ISD) Method for the Measurement of Cyclosporine A, Tacrolimus, Sirolimus/Rapamycin, and Everolimus in Whole Blood Using MassTrak™ Kit.

Cyclosporine A, everolimus, sirolimus, and tacrolimus are the most commonly used immunosuppressant drugs in organ transplant and auto-immune patients. The narrow therapeutic window of these immunosuppressant drugs requires close monitoring of drug blood levels to ensure proper therapeutic response. A quick, robust high-throughput liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was introduced for monitoring whole blood levels of these immunosuppressant drugs with the use of the MassTrak™ Immunosuppressant kit. The assay was carried out in 96-well plate format and requires a simple precipitation step; after which, the supernatant is subjected to liquid chromatography separation (2 min total run time) using a C18 Cartridge column. Identification and quantitation of cyclosporine A, everolimus, sirolimus, and tacrolimus was achieved by employing multiple reaction monitoring (MRM) in positive mode electrospray ionization (ESI). The method exhibits a linear measuring range from 10 to 1500 ng/mL (Cyclosporine A), 1.0-30.0 ng/mL (Everolimus), 1.0-26.0 ng/mL (Sirolimus), and 1.0-30.0 ng/mL (Tacrolimus) and has a within-run and between-run imprecision of <10%.