Evaluation of a rapid micro-scale assay for tacrolimus by liquid chromatography-tandem mass spectrometry
| Autor: | Brian G. Keevil, Donald P. Cooper, M R Morris, Steven J McCann |
|---|---|
| Rok vydání: | 2002 |
| Předmět: |
Chromatography
Formic acid Elution Clinical Biochemistry Enzyme-Linked Immunosorbent Assay General Medicine Mass spectrometry Sensitivity and Specificity Mass Spectrometry Tacrolimus chemistry.chemical_compound chemistry Pharmacokinetics Liquid chromatography–mass spectrometry medicine Humans Regression Analysis Centrifugation Ascomycin Drug Monitoring Ammonium acetate Chromatography High Pressure Liquid Immunosuppressive Agents medicine.drug |
| Zdroj: | Annals of Clinical Biochemistry: International Journal of Laboratory Medicine. 39:487-492 |
| ISSN: | 1758-1001 0004-5632 |
| DOI: | 10.1258/000456302320314502 |
| Popis: | Background: The immunosuppressive drug tacrolimus has complex and unpredictable pharmacokinetics, therefore regular monitoring is required in patients receiving tacrolimus therapy. We have developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for measuring tacrolimus concentrations in whole blood and have compared it with a microparticle enzyme immunoassay. Methods: For the LC-MS/MS assay, samples were prepared in a 96-deep well microtitre plate by adding 10 µL of blood to 40 µL of 0·1 mol/L zinc sulphate solution. Proteins were precipitated by adding 100 µL acetonitrile containing ascomycin internal standard. After vigorous mixing and centrifugation, 20 µL of the supernatant was injected into the LC-MS/MS system. A C18 cartridge (3 mm × 4 mm) was eluted with a step gradient of 50% to 100% methanol containing 2 mmol/L ammonium acetate and 0·1% (v/v) formic acid, at 0·6 mL/min. The column was maintained at 55°C. Results: The retention times were 0·98 min for ascomycin and 0·98 min for tacrolimus. Cycle time was 2·5 min, injection to injection. The analytes were monitored using a Quattro micro tandem mass spectrometer operated in multiple reaction monitoring mode using the following transitions: m/z821 > 768 (tacrolimus) and m/z809 > 756 (ascomycin). The limit of quantitation was 0·5 µg/L and the assay was linear up to 30 µg/L. Precision of the method, over the concentration range 2·5-15·0 µg/L, was < 7% within-batch and < 6% between-batch. Total time to analyse 24 samples including result generation was 90 min. Conclusion: We conclude that the LC-MS/MS method is quick, precise and robust and will provide a fast turn around of results for the transplant physician. |
| Databáze: | OpenAIRE |
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