13c2h4 tacrolimus

Tacrolimus and ammonium acetate were purchased from Sigma‐Aldrich Chemie B.V. (Zwijndrecht, the Netherlands) and ¹³C²H₄‐tacrolimus from Alsa Chim (Illkirch‐Graffenstaden, France). MagSiMUS‐TDM^prep Kit from MagnaMedics containing MagSiMUS‐TDM^prep Type I Particle Mix (Beads), Reconstitution Buffer IS, Organic Precipitation Reagent VI (OPR VI) for methanol‐based protein precipitation and lysis buffer for whole blood were purchased from MagnaMedics Diagnostics B.V. (Geleen, the Netherlands). Water was purified using a MilliPore Advantage A10 System. Methanol and formic acid were purchased from Biosolve BV (Valkenswaard, the Netherlands). Human tacrolimus‐free PBMCs were isolated using Ficoll‐Paque Plus (GE Healthcare Bio‐Sciences AB, Uppsala, Sweden) from buffy coats in citrate blood obtained from Blood bank Sanquin (Rotterdam, the Netherlands). Red blood cell lysis buffer from eBioscience (Affimetrix Inc., San Diego, CA, USA) was used for purification. Cells were counted using a SysmexXOP‐300 cell counter (Sysmex Corporation, Kobe, Japan).

TAC, MPA, and mycophenolic acid-d3 (MPA-d3) which is a deuterated product of mycophenolic acid, were purchased from Toronto Research Chemicals (Toronto, ON, Canada), and [¹³C, ²H₄]-tacrolimus was purchased from Alsachim (Strasbourg, France). Methanol, acetonitrile, and formic acid were obtained from Fisher Chemical (Thermo Scientific, Waltham, MA, USA) at ultra-high performance liquid chromatography/mass spectrometry grades. Ammonium acetate was obtained from Sigma-Aldrich (St. Louis, MO, USA). Ultra-pure water was supplied by the Milli-Q integral water purification system (Merck Millipore, Germany). MagSiMUS-TDM^prep Type I Particle Mix, Organic Precipitation Reagent VI (OPR VI), and Lysis Buffer for whole blood kits were purchased from MagnaMedics Diagnostics B.V. (Geleen, The Netherlands). Ficoll-Paque Plus solution, Dulbecco’s phosphate-buffered saline (D-PBS) without calcium and magnesium ions, and red blood cell lysis buffer were obtained from Solarbio (Beijing, China). PBMCs for the establishment and validation of analytical methods were obtained from healthy volunteers in agreement with the local ethics committee.