Conclusions
Accurate-mass FS-SIM can be successfully utilized for immunosuppressant TDM with good correlation with results generated by standard methods. TurboFlow online SPE allows for a simple "protein crash and shoot" sample preparation protocol. Compared to traditional MRM, analyte quantitation by FS-SIM facilitates a streamlined assay optimization process.
Methods
LC-MS analysis was performed on a TLX-2 multi-channel HPLC with a Q-Exactive Plus mass spectrometer. TurboFlow online SPE was used for sample clean up. The accurate-mass MS was set to positive electrospray ionization mode with FS-SIM for quantitation of tacrolimus, sirolimus, everolimus, and cyclosporine A. MS2 fragmentation pattern was used for compound confirmation.
Results
The method was validated in terms of precision, analytical bias, limit of quantitation, linearity, carryover, sample stability, and interference. Quantitation of tacrolimus, sirolimus, everolimus, and cyclosporine A correlated well with results from an independent reference laboratory (r = 0.926-0.984). Conclusions: Accurate-mass FS-SIM can be successfully utilized for immunosuppressant TDM with good correlation with results generated by standard methods. TurboFlow online SPE allows for a simple "protein crash and shoot" sample preparation protocol. Compared to traditional MRM, analyte quantitation by FS-SIM facilitates a streamlined assay optimization process.