A review of commercially available thrombin generation assays

Abstract

Essentials Overview of the three commercial thrombin generation methods.Description of the sample preparation, data management, and analysis.Description of the similarities and differences in regards to substrates results.Discussion of advantages and disadvantages of the three approaches. Currently there are three commercially available thrombin generation methods. These methods help detect the levels of thrombin generated in patient samples by the use of chromogenic or fluorogenic substrates in plasma or whole blood. Determining the rate of thrombin generation can help indicate if patients are at risk of clotting or bleeding. This review discusses two fluorogenic and one chromogenic method and focuses on similarities and differences of these three methods. The review specifically focuses on the accuracy of commercial substrates used in thrombin generation, and interference that can occur by various plasma proteins, as well as on evaluating the advantages and disadvantages of each method. The commercial chromogenic assay and both fluorogenic assays are able to monitor the rate of thrombin generation and can give indications towards potential coagulation abnormalities. Overall, the main differences between the thrombin generation methods are based on the type of substrate used, sample preparation, and data processing. Despite advancement in this field there are still technical challenges that preclude the widespread use of thrombin generation in clinical applications.

Rivaroxaban pharmacodynamics in healthy volunteers evaluated with thrombin generation and the active protein C system: Modeling and assessing interindividual variability

“…Thrombin generation (TG) has been proposed as an attractive assay to assess in vitro or ex vivo direct FXa inhibitor effect on coagulation based on studies with small series of healthy volunteers or patients, after single or repeated direct Xa inhibitors dose exposure (rivaroxaban, apixaban, edoxaban, or otamixaban). 15–22,26–31 However, some persistent issues remain regarding the most appropriate experimental conditions among different methods, including TF concentrations . Moreover, data regarding the magnitude of TG parameter interindividual variability in subjects receiving direct Xa inhibitors are scarce …”

“…[15][16][17][18][19][20][21][22][26][27][28][29][30][31] However, some persistent issues remain regarding the most appropriate experimental conditions among different methods, including TF concentrations. 32 Moreover, data regarding the magnitude of TG parameter interindividual variability in subjects receiving direct Xa inhibitors are scarce. 15,20,28 In the present study, we sought (a) to characterize plasma TG parameter time profiles in DRIVING healthy volunteers after intake of a 40-mg rivaroxaban single dose, (b) to measure TG under several relevant experimental conditions, and (c) to develop PD models to characterize the relationship between plasma rivaroxaban concentrations and several TG parameters, independently from time of drug intake to estimation of population interindividual variability.…”

Assessing Blood Clotting and Coagulation Factors in Mice

“…However, it also initiates anticoagulant biochemical reactions that gradually cease its production. Assays measuring thrombin generation have existed for many decades (Kintigh, Monagle, & Ignjatovic, 2018). These assays have documented decreased thrombin generation in several coagulation factor deficiencies and in patients using anticoagulant therapy, as well as increased thrombin generation in some congenital and acquired thrombophilias.…”

The effect of pH on thrombin generation–An unrecognized potential source of variation

“…Thrombin generation (TG) assays have become important for determining the global coagulation capacity in plasma. A TG assay was described in 1953, but a more reproducible method was developed and standardized by Hemker et al Calibrated automated thrombin generation (CAT) is widely used to assess TG, but other methods are available with minor differences . With this assay, it is possible to demonstrate a lower capacity in patients prone to bleeding and an increased capacity in patients with a higher risk of thrombosis, and it has been widely used in research to investigate coagulation in various patient groups .…”

“…A TG assay was described in 1953, 1 but a more reproducible method was developed and standardized by Hemker et al [2][3][4] Calibrated automated thrombin generation (CAT) is widely used to assess TG, but other methods are available with minor differences. 5 With this assay, it is possible to demonstrate a lower capacity in patients prone to bleeding and an increased capacity in patients with a higher risk of thrombosis, 4,6,7 and it has been widely used in research to investigate coagulation in various patient groups. 6,7 One drawback of the method has been a rather high imprecision, especially a high interlaboratory variation.…”