Analytical and Bioanalytical Chemistry

, Volume 407, Issue 6, pp 1585–1594 | Cite as

Dried blood spot analysis of creatinine with LC-MS/MS in addition to immunosuppressants analysis

  • Remco A. KosterEmail author
  • Ben Greijdanus
  • Jan-Willem C. Alffenaar
  • Daan J. Touw
Research Paper


In order to monitor creatinine levels or to adjust the dosage of renally excreted or nephrotoxic drugs, the analysis of creatinine in dried blood spots (DBS) could be a useful addition to DBS analysis. We developed a LC-MS/MS method for the analysis of creatinine in the same DBS extract that was used for the analysis of tacrolimus, sirolimus, everolimus, and cyclosporine A in transplant patients with the use of Whatman FTA DMPK-C cards. The method was validated using three different strategies: a seven-point calibration curve using the intercept of the calibration to correct for the natural presence of creatinine in reference samples, a one-point calibration curve at an extremely high concentration in order to diminish the contribution of the natural presence of creatinine, and the use of creatinine-[2H3] with an eight-point calibration curve. The validated range for creatinine was 120 to 480 μmol/L (seven-point calibration curve), 116 to 7000 μmol/L (1-point calibration curve), and 1.00 to 400.0 μmol/L for creatinine-[2H3] (eight-point calibration curve). The precision and accuracy results for all three validations showed a maximum CV of 14.0 % and a maximum bias of −5.9 %. Creatinine in DBS was found stable at ambient temperature and 32 °C for 1 week and at −20 °C for 29 weeks. Good correlations were observed between patient DBS samples and routine enzymatic plasma analysis and showed the capability of the DBS method to be used as an alternative for creatinine plasma measurement.

Graphical Abstract

From blood spot to chromatogram


Dried blood spot Creatinine Tacrolimus Sirolimus Everolimus Cyclosporin A 


Conflict of interest

The authors declare no conflicts of interest.


  1. 1.
    Edelbroek PM, van der Heijden J, Stolk LM (2009) Dried blood spot methods in therapeutic drug monitoring: methods, assays, and pitfalls. Ther Drug Monit 31:327CrossRefGoogle Scholar
  2. 2.
    Vu DH, Alffenaar JW, Edelbroek PM, Brouwers JR, Uges DR (2011) Dried blood spots: a new tool for tuberculosis treatment optimization. Curr Pharm Des 17:2931CrossRefGoogle Scholar
  3. 3.
    Kallner A (2014) Estimated GFR. Comparison of five algorithms: implications for drug dosing. J Clin Pathol 67:609CrossRefGoogle Scholar
  4. 4.
    Biomarkers Definitions Working Group (2001) Biomarkers and surrogate endpoints: preferred definitions and conceptual framework. Clin Pharmacol Ther 69:89CrossRefGoogle Scholar
  5. 5.
    Aburuz S, Millership J, McElnay J (2006) Dried blood spot liquid chromatography assay for therapeutic drug monitoring of metformin. J Chromatogr B Anal Technol Biomed Life Sci 832:202CrossRefGoogle Scholar
  6. 6.
    Lawson G, Cocks E, Tanna S (2013) Bisoprolol, ramipril and simvastatin determination in dried blood spot samples using LC-HRMS for assessing medication adherence. J Pharm Biomed Anal 81–82:99CrossRefGoogle Scholar
  7. 7.
    Koop DR, Bleyle LA, Munar M, Cherala G, Al-Uzri A (2013) Analysis of tacrolimus and creatinine from a single dried blood spot using liquid chromatography tandem mass spectrometry. J Chromatogr B Anal Technol Biomed Life Sci 926:54CrossRefGoogle Scholar
  8. 8.
    Koster RA, Alffenaar JWC, Botma R, Greijdanus B, Touw DJ, Uges DRA, Kosterink JGW (2015 In Press) What is the right blood hematocrit preparation procedure for standards and quality control samples for dried blood spot analysis? BioanalysisGoogle Scholar
  9. 9.
    Koster RA, Alffenaar JW, Greijdanus B, Uges DR (2013) Fast LC-MS/MS analysis of tacrolimus, sirolimus, everolimus and cyclosporin A in dried blood spots and the influence of the hematocrit and immunosuppressant concentration on recovery. Talanta 115:47CrossRefGoogle Scholar
  10. 10.
    Food and Drug Administration, U.S. Department of Health and Human Services (2001) Guidance for Industry, Bioanalytical Method ValidationGoogle Scholar
  11. 11.
    Koster RA, Dijkers EC, Uges DR (2009) Robust, high-throughput LC-MS/MS method for therapeutic drug monitoring of cyclosporine, tacrolimus, everolimus, and sirolimus in whole blood. Ther Drug Monit 31:116CrossRefGoogle Scholar
  12. 12.
    Tan A, Awaiye K, Trabelsi F (2014) Some unnecessary or inadequate common practices in regulated LC-MS bioanalysis. Bioanalysis 6:2751CrossRefGoogle Scholar
  13. 13.
    Clark S, Youngman LD, Palmer A, Parish S, Peto R, Collins R (2003) Stability of plasma analytes after delayed separation of whole blood: implications for epidemiological studies. Int J Epidemiol 32:125CrossRefGoogle Scholar
  14. 14.
    Boyanton BL Jr, Blick KE (2002) Stability studies of twenty-four analytes in human plasma and serum. Clin Chem 48:2242Google Scholar
  15. 15.
    O’Keane MP, Cunningham SK (2006) Evaluation of three different specimen types (serum, plasma lithium heparin and serum gel separator) for analysis of certain analytes: clinical significance of differences in results and efficiency in use. Clin Chem Lab Med 44:662Google Scholar
  16. 16.
    Cannan RK, Shore A (1928) The creatine-creatinine equilibrium. The apparent dissociation constants of creatine and creatinine. Biochem J 22:920Google Scholar
  17. 17.
    Fuller NJ, Elia M (1988) Factors influencing the production of creatinine: implications for the determination and interpretation of urinary creatinine and creatine in man. Clin Chim Acta 175:199CrossRefGoogle Scholar
  18. 18.
    Wyss M, Kaddurah-Daouk R (2000) Creatine and creatinine metabolism. Physiol Rev 80:1107Google Scholar
  19. 19.
    Graves JW (2008) Diagnosis and management of chronic kidney disease. Mayo Clin Proc 83:1064CrossRefGoogle Scholar
  20. 20.
    Timmerman P, White S, Globig S, Ludtke S, Brunet L, Smeraglia J (2011) EBF recommendation on the validation of bioanalytical methods for dried blood spots. Bioanalysis 3:1567CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Remco A. Koster
    • 1
    Email author
  • Ben Greijdanus
    • 1
  • Jan-Willem C. Alffenaar
    • 1
  • Daan J. Touw
    • 1
    • 2
  1. 1.Department of Clinical Pharmacy and Pharmacology, Laboratory for Clinical and Forensic Toxicology and Drugs Analysis, University of GroningenUniversity Medical Center GroningenGroningenThe Netherlands
  2. 2.Department of Pharmacy, Section Pharmacokinetics, Toxicology and TargetingUniversity of GroningenGroningenThe Netherlands

Personalised recommendations