Direct oral anticoagulants (DOACs): From the laboratory point of view

Direct oral anticoagulants (DOACs) represent a new generation of drugs that have been increasingly used in the prevention and treatment of thromboembolic states. According to the mechanism of anticoagulant action, DOACs are divided into two groups: direct inhibitors of thrombin (dabigatran) and direct inhibitors of activated factor X (FXa) (rivaroxaban, apixaban, edoxaban, betrixaban). Compared to the vitamin K antagonists, DOACs are superior in terms of onset of action, pharmacokinetic and pharmacodynamics properties and fixed daily dose without the need for routine coagulation monitoring. Despite these advantages, there are clinical conditions in which laboratory measurement of DOACs should be performed. Although DOACs have an impact on screening haemostasis assays (prothrombin time, PT; activated partial thromboplastin time, aPTT; and thrombin time, TT), these tests are not appropriate for quantifying drug levels. Therefore, specific quantitative methods (LC-MS/MS as a gold standard method for all DOACs, coagulometric and chromogenic assays for dabigatran, and chromogenic anti-Xa assays with drug-specific calibrators for inhibitors of FXa) should only be used for determination of DOACs concentration. The aim of this review is to present all aspects of laboratory assessment of DOACs, including pre-analytical, analytical and post-analytical factors in the overall testing process with a special accent on the available specific quantitative methods for measurement of DOACs in circulation

Analytical verification and comparison of chromogenic assays for dabigatran, rivaroxaban and apixaban determination on BCSXP and STA Compact Max analysers

Introduction: The aim of the study was to perform analytical verification and comparison of chromogenic assays for determination of dabigatran, rivaroxaban and apixaban concentration on BCSXP and STA Compact Max analysers. Materials and methods: Precision, linearity, measurement uncertainty estimation and determination of limit of blank, limit of determination and limit of quantification were calculated. Analytical performance specifications were set according to manufacturer specifications and literature data on between laboratory variability. Comparison of the methods was done using Bland-Altman and Passing-Bablok regression analysis. Results: Obtained results have shown acceptable precision on STA Compact Max only for dabigatran (CV = 3.5%) at lower concentration level comparing to manufacturer declaration (CV = 3.6%). On BCSXP, the highest coefficient of variation has been shown for apixaban (6.1%) at lower concentration level. Within laboratory precision was not met on STA Compact Max for all assays. Bland-Altman analysis has shown statistically significant bias for dabigatran (23.2%, 95%CI 11.2 – 35.3; P < 0.001) and apixaban (8.4%, 95%CI 1.2 – 15.6; P = 0.023). Passing-Bablok regression analysis has shown systematic and proportional deviation between methods for rivaroxaban (y = 6.52 (2.94 to 11.83) + 0.84 (0.80 to 0.89) x. Conclusion: Chromogenic assays for dabigatran, rivaroxaban and apixaban on BCSXP and STA Compact Max analysers are shown as methods with satisfactory long-term analytical performance specifications for determination of direct oral anticoagulants in clinical laboratories. However, we cannot recommend interchangeable use because of the significant bias between assays

Croatian Society of Medical Biochemistry and Laboratory Medicine: National recommendations for blood collection, processing, performance and reporting of results for coagulation screening assays prothrombin time, activated partial thromboplastin time, thrombin time, fibrinogen and D-dimer

A modern diagnostic laboratory offers wide spectrum of coagulation assays utilized in the diagnosis and management of patients with haemostatic disorders, preoperative screening and anticoagulation therapy monitoring. The recent survey conducted among Croatian medical biochemistry and transfusion laboratories showed the existence of different practice policies in particular phases of laboratory process during coagulation testing and highlighted areas that need improvement. Lack of assay standardization together with non-harmonized test results between different measurement methods, can potentially lead to incorrect decisions in patient’s treatment. Consequently, patient safety could be compromised. Therefore, recommended procedures related to preanalytical, analytical and postanalytical phases of prothrombin time, activated partial thromboplastin time, thrombin time, fibrinogen and D-dimer testing are provided in this review, aiming to help laboratories to generate accurate and reliable test results

Interference of M-protein on prothrombin time test – case report.

The aim of this report was to present a case of interference on prothrombin time (PT) test that directed further laboratory diagnostics and resulted with final detection of monoclonal gammopathy in an 88-year old man. Routine coagulation testing during medical examination at Emergency Department revealed unmeasurable PT (< 7% activity) and activated partial thromboplastin time (aPTT) within reference range. After repeated sampling for coagulation testing, PT was unmeasurable again, as well as fibrinogen level (< 0.8 g/L), thrombin time (TT) was significantly prolonged (107 seconds) and aPTT was within reference range. In both plasma samples refrigerated at 4 °C overnight, white gelatinous precipitate was visible between the cell and plasma layers and the presence of monoclonal protein (M-protein) was suggested in our patient. Further laboratory diagnostics revealed total serum proteins at concentration of 123 g/L and the presence of M-protein IgG lambda (?) at concentration of 47.1 g/L. These results suggested monoclonal gammopathy as an underlying pathophysiological condition in our patient. Activities of coagulation factors II, V, VII and X were within reference ranges or increased. These results and correction of unmeasurable PT result to 67% in mixing test with commercial normal plasma suggest in vitro rather than in vivo interference of M-protein on PT result. In contrast, significantly prolonged TT results in all analysed samples suggest impact of M-protein on this global coagulation test due to possible effect on fibrin polymerization

Reporting of activated partial thromboplastin time (aPTT): Could we achieve better comparability of the results?

Activated partial thromboplastin time (aPTT) is determined and reported as clotting time in seconds aPTT(s), but it is presumed that reporting results as patient-to-normal clotting time ratio, aPTT(r), could minimize within-laboratory variability. The aim of study was to investigate differences in reporting aPTT results that can affect comparability of the results among Croatian laboratories and suggest further steps for its harmonization. The questionnaire on aPTT reporting practice was distributed to 83 laboratories through Survey Monkey application in March 2019 as the part of the first regular round of Croatian Centre for Quality Assessment in Laboratory Medicine proficiency testing. The survey response rate was 0.49. Majority of laboratories report aPTT results as both, seconds and ratio. Participants reported use of 23 different aPTT(s) reference intervals along with 17 different combinations of reagent/coagulometer and 25 aPTT(s) denominators of different origin for aPTT(r) calculation. Despite the same aPTT(s) results, the use of different denominators caused a dispersion of aPTT(r) results that can lead to exceeding external quality assessment performance criteria of 7%, particularly when results were compared for the same reagent group only. By applying aPTT(s) reference interval mean as denominator for calculation of aPTT(r) reference interval better concordance to harmonized one was obtained (17 vs. 27; χ2 = 3.972; P = 0.046). In order to improve comparability of the results, laboratories are advised to use mean of aPTT(s) reference interval as denominator for aPTT(r) calculation. Type of coagulometer need to be considered when evaluating aPTT proficiency test results and its currently acceptable limit of performance evaluated accordingly

Thromboembolism in a 14-year-old boy – case report

Uvod: Opisuje se slučaj venske tromboembolije (VTE) u 14-godišnjeg dječaka u kojega se razvila duboka venska tromboza desne noge, a potom i plućna embolija komplicirana plućnim infarktom. Cilj: Prikazati rezultate laboratorijske dijagnostike koji upućuju na mogući uzrok nastanka VTE u ovog bolesnika. Materijali i metode: Laboratorijska obrada VTE uključila je određivanje antifosfolipidnih antitijela: lupus antikoagulanta (LAC) i antikardiolipinskih antitijela (ACA) klase IgG i IgM, inhibitora zgrušavanja (antitrombin, protein C, protein S) i fibrinolize (inhibitor aktivatora plazminogena-1 (PAI-1)), određivanje rezistencije na aktivirani protein C (APCR) te molekularnu dijagnostiku čimbenika tromboembolije: genotipizacija polimorfizama u genu za PAI-1 i metilentetrahidrofolat reduktazu (MTHFR), mutacije G20210A faktora II zgrušavanja (protrombina) i mutacije G1691A faktora V zgrušavanja (faktor V Leiden). Rezultati: U bolesnika su utvrđena antifosfolipidna antitijela: lupus antikoagulant (omjeri LAC 3,0 i 2,76) i antikardiolipinska antitijela klase IgG (104,7 i 103,7 GPL-U/mL) u dva nezavisna mjerenja u razmaku od devet tjedana. Ovi rezultati uz kliničku sliku ukazuju na prisutnost antifosfolipidnog sindroma. Molekularnom dijagnostikom utvrđen je polimorfizam gena za PAI-1 (geno-tip 4G/5G), što je u skladu s izmjerenom povećanom koncentracijom PAI-1 u plazmi (4,9 IU/mL). Zaključak: Laboratorijskom dijagnostikom tromboembolije u opisanog bolesnika utvrđen je antifosfolipidni sindrom kao čimbenik rizika za nastanak VTE. Iako se polimorfizam PAI-1 ne smatra neovisnim čimbenikom rizika za nastanak VTE, moguće je da u bolesnika s utvrđenim čimbenikom tromboembolije kao što je antifosfolipidni sindrom i ovaj polimorfizam ima ulogu u nastanku VTE. Infekcija i mirovanje koji su prethodili trombozi mogli bi biti dodatni čimbenici rizika za nastanak VTE u ovog bolesnika. Prikazani slučaj govori u prilog dosadašnjim saznanjima da je pojava VTE najčešće rezultat međudjelovanja genetskih i stečenih čimbenika rizika.Introduction: A case of venous thromboembolism (VTE) in a 14-year-old boy who developed deep venous thrombosis of the right leg followed by pulmonary embolism complicated with pulmonary infarction is described in this article. Aim: To present results of laboratory diagnosis suggesting the possible cause of VTE occurrence in this patient. Materials and methods: Laboratory diagnosis of VTE included determination of antiphospholipid antibodies: lupus anticoagulant (LAC) and anticardiolipin antibodies (ACA) IgG and IgM classes, inhibitors of coagulation (antithrombin, protein C, protein S) and fibrinolysis (plasminogen activator inhibitor-1 (PAI-1)), determination of activated protein C resistance (APCR) and molecular diagnosis of thrombophilic factors: genotyping polymorphisms in the gene for PAI-1 and methylenetetrahydrofolate reductase (MTHFR), mutation G20210A for coagulation factor II (prothrombin) and mutation G1691A for coagulation factor V (factor V Leiden). Results: The following antiphospholipid antibodies were demonstrated in the patient: lupus anticoagulant (LAC ratios 3.0 and 2.76) and anticardiolipin antibodies of IgG class (104.7 and 103.7 GPL-U/mL) on two independent measurements nine weeks apart. These results, along with clinical presentation, suggested the presence of antiphospholipid syndrome. Molecular diagnosis confirmed polymorphism in the gene for PAI-1 (genotype 4G/5G), which was in accordance with elevated concentration of PAI-1 measured in plasma (4.9 IU/mL). Conclusion: In this patient, laboratory diagnosis of thrombophilia revealed antiphospholipid syndrome as a risk factor for VTE. Although PAI-1 polymorphism is not considered as an independent risk factor for VTE, it is possible that in patients with established thrombophilic factor such as antiphospholipid syndrome this polymorphism may play a role in VTE occurrence. Infection and prolonged bed rest that preceded thrombosis could have been additional risk factors for VTE occurrence in this patient. The reported case supports the current concepts according to which VTE occurrence most frequently results from interaction of genetic and acquired risk factors

Algorithm for Rapid Exclusion of Clinically Relevant Plasma Levels of Direct Oral Anticoagulants in Patients Using the DOAC Dipstick:An expert consensus paper

With the widespread use of direct oral anticoagulants (DOACs), there is an urgent need for a rapid assay to exclude clinically relevant plasma levels. Accurate and rapid determination of DOAC levels would guide medical decision-making to (a) determine the potential contribution of the DOAC to spontaneous or trauma-induced hemorrhage; (b) identify appropriate candidates for reversal, or (c) optimize the timing of urgent surgery or intervention. The DOAC Dipstick test uses a disposable strip to identify factor Xa- or thrombin inhibitors in a urine sample. Based on the results of a systematic literature search followed by an analysis of a simple pooling of five retrieved clinical studies, the test strip has a high sensitivity and an acceptably high negative predictive value when compared with levels measured with liquid chromatography tandem mass spectrometry or calibrated chromogenic assays to reliably exclude plasma DOAC concentrations &gt;30ng/mL. Based on these data, a simple algorithm is proposed to enhance medical decision-making in acute care indications useful primarily in hospitals not having readily available quantitative tests and 24/7. This algorithm not only determines DOAC exposure but also differentiates between factor Xa- and thrombin inhibitors to better guide clinical management.</p

Security of information systems in state administration bodies

U današnje vrijeme tijela državne uprave raspolažu s visokim stupnjem tajnosti kvalificiranih i nekvalificiranih podataka koji mogu biti predmet interesa raznih obavještajnih službi, gospodarskih subjekata, organizacija, ali i kriminalnih i terorističkih skupina. U osnovi sigurnosti svakog informacijskih sustava je osiguravanje takvih informacija, održavanje povjerljivosti, integriteta i dostupnosti (CIA) informacija, osiguravajući da informacije ne budu ugrožene na bilo koji način. Usprkos tome, sigurnosne prijetnje, incidenti, ranjivosti i rizici problem su u mnogim državnim tijelima, a jedan od glavnih uzroka ovog problema je slabo razumijevanje ključnih sigurnosnih i propisanih čimbenika. Zbog toga možemo reći da je sigurnost informacijskih sustava neizostavan dio poslovanja i uspješnosti državnih institucija u Republici Hrvatskoj, posebice onih koje koriste modernu i visoku tehnologiju. Cilj ovog rada je prikazati sigurnosne, propisane aspekte i važnost pravilnog funkcioniranja sigurnosti informacijskih sustava u tijelima državne uprave u Republici Hrvatskoj.Abstract Nowadays, state administration bodies have a high degree of secrecy for qualified and unqualified data that may be the subject of interest of various intelligence services, economic entities, organizations, but also criminal and terrorist groups. The main task of the security of any information system is the provision of such information, maintaining the confidentiality, integrity and availability (CIA) of information, ensuring that informations aren't compromised in any way. Nevertheless, security threats, incidents, vulnerabilities, and risks are a problem in many government bodies, and one of the main causes of this problem is a poor understanding of key security and regulatory factors. Therefore, we can say that the security of information systems is an indispensable part of the business and success of all state institutions in the Republic of Croatia, especially those that use modern and high technology. The main goal of this paper is to present the security, prescribed aspects and the importance of the proper functioning of the security information systems in state administration bodies in the Republic of Croatia

Security of information systems in state administration bodies

U današnje vrijeme tijela državne uprave raspolažu s visokim stupnjem tajnosti kvalificiranih i nekvalificiranih podataka koji mogu biti predmet interesa raznih obavještajnih službi, gospodarskih subjekata, organizacija, ali i kriminalnih i terorističkih skupina. U osnovi sigurnosti svakog informacijskih sustava je osiguravanje takvih informacija, održavanje povjerljivosti, integriteta i dostupnosti (CIA) informacija, osiguravajući da informacije ne budu ugrožene na bilo koji način. Usprkos tome, sigurnosne prijetnje, incidenti, ranjivosti i rizici problem su u mnogim državnim tijelima, a jedan od glavnih uzroka ovog problema je slabo razumijevanje ključnih sigurnosnih i propisanih čimbenika. Zbog toga možemo reći da je sigurnost informacijskih sustava neizostavan dio poslovanja i uspješnosti državnih institucija u Republici Hrvatskoj, posebice onih koje koriste modernu i visoku tehnologiju. Cilj ovog rada je prikazati sigurnosne, propisane aspekte i važnost pravilnog funkcioniranja sigurnosti informacijskih sustava u tijelima državne uprave u Republici Hrvatskoj.Abstract Nowadays, state administration bodies have a high degree of secrecy for qualified and unqualified data that may be the subject of interest of various intelligence services, economic entities, organizations, but also criminal and terrorist groups. The main task of the security of any information system is the provision of such information, maintaining the confidentiality, integrity and availability (CIA) of information, ensuring that informations aren't compromised in any way. Nevertheless, security threats, incidents, vulnerabilities, and risks are a problem in many government bodies, and one of the main causes of this problem is a poor understanding of key security and regulatory factors. Therefore, we can say that the security of information systems is an indispensable part of the business and success of all state institutions in the Republic of Croatia, especially those that use modern and high technology. The main goal of this paper is to present the security, prescribed aspects and the importance of the proper functioning of the security information systems in state administration bodies in the Republic of Croatia

Development and application of e trade in Republic of Croatia

Cilj ovog rada jest prikazati razvoj elektroničke trgovine u Republici Hrvatskoj te utvrditi razinu njene primjene u današnjem poslovanju. Uz navedeno, u radu će se identificirati izazovi primjene elektroničke trgovine, načini prodaje te ključne razvojne smjernice u budućnosti