Etablierung eines automatisierten Verfahrens zur Quantifizierung von Plasmakonzentrationen direkter oraler Antikoagulantien
by Anna Abratis
Date of Examination:2023-10-18
Date of issue:2023-10-12
Advisor:PD Dr. Dr. Moritz Thomas Schnelle
Referee:Prof. Dr. Manuel Wallbach
Referee:Prof. Dr. Margarete Schön
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EnglishMore than a decade after their first approval, the imageability of direct oral anticoagulants (DOACs) still poses challenges to laboratories worldwide. Even though calibrator- and matrix-dependent chromogenic tests are now quite common, they reach their limits especially in emergency situations with limited information on the ingested drug. LCMS-based measurement methods, the current gold standard in DOAC determination, bear potential in this regard. However, due to the need for trained personnel, long runtimes and limited availability, their use today is mainly limited to scientific questions. We coupled an LCMS-based DOAC determination to an automated sample preparation module (CLAM-2030, Shimadzu) and evaluated the practicality of the 24/7 routine in this constellation. Main objectives: (1) to compare measured plasma concentrations with the LCMS-8050 system coupled to the CLAM-2030 (both Shimadzu) with results of commonly used chromogenic assays (DTI assay and anti-Xa assay, HemosIL) (2) to evaluate the practicability of the automated measurement methodology, considering the information gain. Methodology: Quantification of DOACs in 56 patient samples on an LCMS system with an automated sample preparation module. Performance of the measurement separated according to the active ingredients (apixaban, rivaroxaban and dabigatran). Automated protein precipitation and separation of samples by filtration using CLAM-2030, which also functions as an autosampler for the connected LCMS. Comparison of plasma concentrations with the results of Anti-Xa (apixaban, rivaroxaban) and DTI assay (dabigatran). Results: (1) Good correlation in method comparison for all three analytes (apixaban r=0.984, rivaroxaban r=0.986, dabigatran r=0.988). Compliance with the precision specified by the manufacturer for calibrations and control measurements (concentration ranges from about 10 to 500ng/ml). (2) Easy handling of the automated system after a short training period. Average measurement time of about 6 minutes. Only minor interferences between the individual parameters, which could be clearly assigned to technical noise. Conclusion: The described combination of LCMS and an automated sample preparation module convinces with easy handling and fast sample run times. The transition of use cases for LCMS-based DOAC determination from purely scientific questions to the use in (24/7) routine measurements becomes conceivable. Compared to previous measurement methods, the procedure described offers the possibility of quantifying all DOACs in a single measurement run. This bears enormous potential for better assessing the coagulation of patients in different clinical (emergency) settings.
Keywords: DOAC; LCMS; TDM; anticoagulants; quantification