Rapid simultaneous quantification of immunosuppressants in transplant patients by turbulent flow chromatography combined with tandem mass spectrometry.
BACKGROUND
Immunosuppressant therapeutic drug monitoring (TDM) is an important requirement in the management of post-transplant patients. Our aim was to develop and evaluate a robust high-throughput method using turbulent flow chromatography (TFC) coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the simultaneous quantification of cyclosporin A (CsA), tacrolimus (FK 506) and sirolimus.


METHODS
A total of 1483 EDTA-blood pre-dosage samples from 147 kidney, 67 liver, 15 kidney/pancreas, and 48 bone marrow recipients were collected. After hemolysis and protein precipitation of 50 microl blood, fast and efficient on-line matrix elimination was achieved using turbulent flow chromatography. Tandem mass spectrometric detection and quantification was performed using multiple reaction monitoring (MRM).


RESULTS
The total analysis time of the column switching method was 3 min. The method was linear from 4.5 to 1500 ng/ml for cyclosporin A, from 0.2 to 100 ng/ml for tacrolimus, and from 0.4 to 100 ng/ml for sirolimus. The accuracy was >95%. Within and between-run assay variation coefficients ranged from 2.4% to 9.3%. Excellent correlation with other standard methods (immunoassay, HPLC) was observed.


CONCLUSIONS
The presented turbulent flow chromatography-tandem mass spectrometric platform offers a very fast, simple and economical method with an excellent validation profile and is well suited for daily pre- and post-dosage immunosuppressant monitoring.
Ceglarek
Uta
U
Institute of Laboratory Medicine, Clinical Chemistry and Molecular Diagnostics, University Hospital Leipzig, Liebigstrasse 27 D-04103 Leipzig, Germany.
Lembcke
Jan
J
Fiedler
Georg Martin
GM
Werner
Maria
M
Witzigmann
Helmut
H
Hauss
Johann Peter
JP
Thiery
Joachim
J
D016026:Bone Marrow Transplantation; D002624:Chemistry, Clinical / Q000379:methods; D016572:Cyclosporine / Q000097:blood; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D007166:Immunosuppressive Agents / Q000097:blood*; D016030:Kidney Transplantation; D016035:Pancreas Transplantation; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D020123:Sirolimus / Q000097:blood; D016559:Tacrolimus / Q000097:blood
D007166:Immunosuppressive Agents; D016572:Cyclosporine; D020123:Sirolimus; D016559:Tacrolimus