The Use of Fluorogenic Substrates to Monitor Thrombin Generation for the Analysis of Plasma and Whole Blood Coagulation

Ramjee, Manoj K.

doi:10.1006/abio.1999.4380

Cited by 41 publications

(48 citation statements)

References 15 publications

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“…Thrombin generation in the INCA is measured in international units 1,2 and not in coagulation seconds, which is sometimes difficult to standardize. 3,4 In the important ascending branch of thrombin generation, the INCA is extremely sensitive to low-grade contact activation that may often occur in patients.…”

Section: Intrinsic Coagulation Activity Assay In Individual and Poolementioning

confidence: 99%

Intrinsic Hemostasis Activation by Freezing and Thawing of Plasma

Stief

Otto

Renz

2007

Clin Appl Thromb Hemost

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Low-grade contact activation of hemostasis is clinically relevant. Freezing/thawing of plasma was studied in the intrinsic coagulation activity assay. Normal plasmas were frozen at −80°C or −20°C and thawed at 37°C or 23°C. These plasmas and unfrozen samples were activated by SiO2 -CaCl2. Freezing/thawing of normal plasma induced about 100-fold more thrombin activity at 5 minutes coagulation reaction time than the respective unfrozen samples. Freezing at −80°C induces more artificial changes than freezing at −20°C. In 9 of 10 plasmas of patients receiving coumarin, nearly no additional thrombin is generated within a 12-minute coagulation reaction time. Minor procoagulant changes of plasma might be dangerous in patients with insufficient liver function, who might not tolerate a therapy with fresh frozen plasma, which behaves as a procoagulant because of its matrix changes. The intrinsic coagulation activity assay allows the measurement of low-grade contact activation of frozen/thawed plasma.

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Section: Intrinsic Coagulation Activity Assay In Individual and Poolementioning

confidence: 99%

Intrinsic Hemostasis Activation by Freezing and Thawing of Plasma

Stief

Otto

Renz

2007

Clin Appl Thromb Hemost

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“…The use of chromogenic substrates requires measuring optical density, which renders whole blood and platelet rich plasma samples problematic for colorimetric measurements [16]. Fibrinogen clotting results in turbidity of plasma samples which interferes negatively with absorbance readings [16][17][18].…”

Section: Introductionmentioning

confidence: 99%

“…The potential advantages of fluorogenic assays over chromogenic assays include the ability to use a range of sample types such as platelet poor (PPP), platelet rich (PRP) and whole blood samples, as fluorescence is not as influenced by the opacity of the sample as absorbance [18]. Although fluorogenic assays to assess thrombin generation have been developed, there is accumulating evidence that FXa may represent a better target as it occupies a critical junction in the coagulation cascade [20].…”

mentioning

confidence: 99%

Development of a fluorescent anti-factor Xa assay to monitor unfractionated and low molecular weight heparins

Harris

Castro-López

Hammadi

et al. 2010

Talanta

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Fluorogenic assays have many potential advantages over traditional clot-based and chromogenic assays such as the absence of interference from a range of factor deficiencies as well as offering the possibility of assays in platelet rich plasma or whole blood. A fluorogenic anti-factor Xa (anti-FXa) assay has been developed for the determination of heparin-like anticoagulants including unfractionated heparin (UFH), low-molecular weight heparins (LMWHs), namely enoxaparin and tinzaparin, and the synthetic heparinoid danaparoid, in commercial human pooled plasma. The assay was based on the complexation of heparinspiked plasmas with exogenous FXa at a concentration of 4 nM in the presence of 0.9 µM of the fluorogenic substrate methylsulfonyl-D-cyclohexylalanyl-glycyl-arginine-7-amino-4-methylcoumarin acetate (Pefafluor FXa). Pooled plasma samples were spiked with concentrations of anticoagulants in the range 0 to 1.6 U/ml. The assay was capable of the measurement of UFH and danaparoid in the range 0-1 U/ml, and enoxaparin and tinzaparin in the range 0-0.8 U/ml and 0-0.6 U/ml, respectively. Assay percentage coefficients of variation were typically below 7 %.3

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“…This expectation appears justified, as the thrombin generation test has since improved significantly by the introduction of continuous techniques employing chromogenic and fluorogenic substrates [7][8][9][10], thus replacing the original subsampling techniques which are laborious and time consuming. Simultaneous handling of many samples is possible by this method, thus saving time and costs.…”

Section: Introductionmentioning

confidence: 99%

Calibrating Thrombin Generation in Different Samples: Less Effort with a Less Efficient Substrate

Tapp¹,

Grundmann²,

Kusch³

et al. 2009

TOATHERTJ

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Abstract:The technique of thrombin generation has been established as a promising tool for the diagnosis, risk estimation, and perhaps prognosis of coagulation insufficiencies. Without further experimentation an indication can be obtained if a sample donor carries the risk for haemophilia or thrombophilia, either congenital or acquired. A comparison of two different thrombin generation assays is presented, demonstrating that a currently not commercially available test allows an easier and cheaper calibration than its commercial counterpart. This is achieved by employment of a less efficient but highly specific peptide substrate for thrombin and the involvement of low amounts of analyte (plasma samples).

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The Use of Fluorogenic Substrates to Monitor Thrombin Generation for the Analysis of Plasma and Whole Blood Coagulation

Cited by 41 publications

References 15 publications

Intrinsic Hemostasis Activation by Freezing and Thawing of Plasma

Intrinsic Hemostasis Activation by Freezing and Thawing of Plasma

Development of a fluorescent anti-factor Xa assay to monitor unfractionated and low molecular weight heparins

Calibrating Thrombin Generation in Different Samples: Less Effort with a Less Efficient Substrate

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