A rapid, sensitive and specific assay to measure TF activity based on chromogenic determination of thrombin generation

Abstract A new specific assay for urinary creatinine is described. The procedure is based on the separation of creatinine from other urinary components by Sephadex gel filtration, and subsequent measurement of the compound at 235 nm against a phosphate buffer blank. No interference by other substances has been encountered. The standard deviations in a series of assays using standard solutions are less than 2%, and the recovery of added known amounts of creatinine from urine is excellent. The method has been found satisfactory for simultaneous determination of creatinine and hippuric acid in urine.

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Effect of substrate and fibrin polymerization inhibitor on determination of plasma thrombin generation in vitro

Thrombosis Research ◽

10.1016/j.thromres.2006.05.008 ◽

2007 ◽

Vol 119 (6) ◽

pp. 667-677 ◽

Cited By ~ 13

Author(s):

A. Lau ◽

L.R. Berry ◽

L.G. Mitchell ◽

A.K.C. Chan

Keyword(s):

Thrombin Generation ◽

Fibrin Polymerization ◽

Polymerization Inhibitor

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Thrombin generation and venous thromboembolism

Hämostaseologie ◽

10.1055/s-0037-1616919 ◽

2008 ◽

Vol 28 (01/02) ◽

pp. 37-39 ◽

Cited By ~ 6

Author(s):

S. Eichinger

Keyword(s):

At Risk ◽

Venous Thromboembolism ◽

Thrombin Generation ◽

Recurrence Risk ◽

Hormonal Contraceptives ◽

Antithrombin Deficiency ◽

Unknown Risk ◽

Patients At Risk ◽

Recurrent Venous Thrombosis

SummaryVenous thromboembolism is a chronic and potential fatal disease. Determination of recurrence risk is time-consuming and costly, and sometimes not feasible: many patients carry more than one risk factor, the relevance of some factors with regard to risk of recurrence is unknown, and existence of thus far unknown risk factors must be considered. A laboratory assay that measures multifactorial thrombophilia would be useful to identify patients at risk of thrombosis. The process of thrombin generation is the central event of the hemostatic process. Thrombin generation is increased in patients at risk of thrombosis including those with antithrombin deficiency or those who are taking hormonal contraceptives. Risk of first and recurrent venous thrombosis is higher in patients with increased thrombin generation. Thus, by use of a simple global marker of coagulation stratification of patients according to their risk of thrombosis is possible. Future studies are needed to improve the management of patients with VTE and increased thrombin generation.

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A Rapid and Specific Assay for the Determination of Lamotrigine in Human Plasma by Normal-Phase HPLC

Therapeutic Drug Monitoring ◽

10.1097/00007691-199702000-00020 ◽

1997 ◽

Vol 19 (1) ◽

pp. 100-107 ◽

Cited By ~ 20

Author(s):

Anna Bartoli ◽

Roberto Marchiselli ◽

Giuliana Gatti

Keyword(s):

Human Plasma ◽

Normal Phase ◽

Specific Assay ◽

Normal Phase Hplc

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Determination of platelet factor 3 by thrombin generation using a micro-coagulation assay

Thrombosis Research ◽

10.1016/0049-3848(82)90077-9 ◽

1982 ◽

Vol 27 (3) ◽

pp. 303-310 ◽

Cited By ~ 3

Author(s):

B. Kirchhof ◽

L. Balleisen

Keyword(s):

Thrombin Generation ◽

Platelet Factor ◽

Coagulation Assay

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Development of a Fluorescent-Bead-Based Multiplex Immunoassay To Determine Immunoglobulin G Subclass Responses to Neisseria meningitidis Serogroup A and C Polysaccharides

Clinical and Vaccine Immunology ◽

10.1128/cvi.00478-07 ◽

2008 ◽

Vol 15 (8) ◽

pp. 1188-1193 ◽

Cited By ~ 38

Author(s):

Richarda M. de Voer ◽

Fiona R. M. van der Klis ◽

Carla W. A. M. Engels ◽

Ger T. Rijkers ◽

Elisabeth A. Sanders ◽

...

Keyword(s):

Neisseria Meningitidis ◽

Immunoglobulin G ◽

Serum Immunoglobulin ◽

Igg Subclass ◽

Fluorescent Particle ◽

Multiplex Immunoassay ◽

Specific Assay ◽

Flow Cytometric ◽

Immunoglobulin G Subclass

ABSTRACT A fluorescent-particle-based multiplex flow cytometric immunoassay (MIA) for the detection of serum immunoglobulin G (IgG) and two IgG subclasses, IgG1 and IgG2, specific for Neisseria meningitidis serogroup A (MenA) and C (MenC) polysaccharides (PS) was developed. The assay comprised three separate duplex assays, one for the detection of the IgG response to MenA and MenC PS, another for the detection of the IgG1 response to MenA and MenC PS, and a third for the detection of the IgG2 response to MenA and MenC PS. Next, the three separate duplex assays were combined and analyzed as a hexaplex assay. No interference between monoplex, duplex, and hexaplex assays was observed, and the assay was found to have low intra- and interassay variation (<9.0% and <27%, respectively). Comparison of the meningococcal subclass MIA to the in-house enzyme-linked inmmunosorbent assays showed a good correlation (R ≥ 0.85) for each of the subclasses. We conclude that the hexaplex meningococcal subclass MIA is an easy and specific assay for the determination of anti-MenA and anti-MenC PS subclass IgG, requiring minimal amounts of serum to study IgG subclass responses to vaccines.

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The Routine Determination of the Endogenous Thrombin Potential, First Results in Different Forms of Hyper- and Hypocoagulability

Thrombosis and Haemostasis ◽

10.1055/s-0038-1656024 ◽

1997 ◽

Vol 77 (04) ◽

pp. 629-636 ◽

Cited By ~ 88

Author(s):

Simone Wielders ◽

Manjari Mukherjee ◽

Jan Michiels ◽

Dirk T S Rijkers ◽

Jean-Pierre Cambus ◽

...

Keyword(s):

Thrombin Generation ◽

Antithrombin Deficiency ◽

Endogenous Thrombin Potential ◽

Vein Thrombosis ◽

Heparin Administration ◽

First Results ◽

Artery Disease ◽

Deep Vein ◽

Factor Concentration

SummaryThe area under the thrombin generation curve (the endogenous thrombin potential; ETP) has been proposed as a parameter for plasma-based hypercoagulability and to monitor anticoagulant treatment. We present an ETP assay for the routine laboratory using a centrifugal analyser. Throughput is 30 samples/h, within and between run imprecision is 4-5.6%. Suitable substrates were developed for the ranges of 10-500% and 2-100% of normal.Independent of tissue factor concentration (if >4 pM), the normal value of the extrinsic ETP is 384.8 ±51.7 nM.min. The intrinsic ETP, triggered by ellagic acid, is 414 ± 41 nM.min.The ETP is decreased to 15 and 35% of normal by oral anticoagulation (INR 2.5-4.0) and by heparin administration (APTT 1.5-2.5 X control).The ETP is increased in untreated subjects with congenital antithrombin deficiency and in women using oral contraceptives. In deep vein thrombosis (phlebographically confirmed), it is increased by 29.4% (extrinsic) and 53% (intrinsic). In (angiographically assessed) coronary artery disease the increase is by 10% and 17% respectively.

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Comparison of a New Automated Amidolytic Assay of Thrombin Generation with “ Prothrombin Time” and “Thrombotest”.

10.1055/s-0039-1687509 ◽

1979 ◽

Author(s):

J.J.C. Jonker ◽

L.H.M. van Riel ◽

M.M.P. Paulssen

Keyword(s):

Prothrombin Time ◽

Whole Blood ◽

Thrombin Generation ◽

Chromogenic Substrate ◽

Automated Method

An automated method was developed for the determination of thrombin generation in citrated whole blood during activation by thromboplastin. The thrombin generated was allowed to split the chromogenic substrate Tos-Gly-Pro-Arg-pNA yielding p-Nitroaniline. The comparison with this method and “Prothrombin time” in 50 samples was significant (r = 0,88 P < 0,001) and also in 503 samples with “Thrombotest” (r = 0,73 P < 0,001). This assay may serve as a specific, precise and fast and cheaper alternative for “prothrombin time” and “Thrombotest” in large Thrombosis Services.

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Routine Measurement of Endogenous Thrombin Potential (ETP) To Assess Hemostatic Function in Patients with Liver Cirrhosis.

Blood ◽

10.1182/blood.v108.11.4014.4014 ◽

2006 ◽

Vol 108 (11) ◽

pp. 4014-4014

Author(s):

Alain P. Gadisseur ◽

Jeoffrey Schouten ◽

Sven Franque ◽

Marc Van der Planken ◽

Peter Michielsen ◽

...

Keyword(s):

Liver Cirrhosis ◽

Thrombin Generation ◽

Pearson Correlation ◽

Area Under The Curve ◽

Classification Systems ◽

Bleeding Tendency ◽

Endogenous Thrombin Potential ◽

Automated Assay ◽

The Mean

Abstract The maintenance of hemostasis through the production of most of the coagulation proteins is a basic liver function. In patients with cirrhosis of the liver the decrease in these proteins is one of the contributory factors to an increased bleeding tendency. Normally the hemostatic capacity of the liver is measured through routine clotting tests as the activated Partial Thromboplastin Time (aPTT) and Prothrombin Time (PT). In the Child classification for liver cirrhosis the coagulation as expressed by the PT is one of the determinants. Recently a test has become available which will make it possible to routinely measure the endogenous thrombin generation potential (ETP) which may be a better alternative. In this test according to the method first described by Hemker (1993) thrombin generation is continuously measured by use of a chromogenic substrate. Results are calculated as area under the curve and as a percentage of normal. We analysed 110 patients with liver cirrhosis classified according to the Child classification, 79 patients with stage A, 19 stage B and 12 stage C, without known pre-existing coagulation abnormalities like inherited bleeding disorders, or anticoagulant drugs. In these patients ETP, APTT, PT/INR, FV, FVII and FXI were measured. We used a fully automated assay for the determination of the endogenous thrombin potential (ETP) on the BCS® System (both Dade Behring, Marburg, Germany). The results for the mean PT were 81% (CI95 77 - 85) for Child A patients, 53% (CI95 44 - 63) for Child B, and 41% (CI95 33 - 50) for Child C. For the mean INR the results were 1.11 (CI95 1.08 - 1.14) for Child A patients, 1.65 (CI95 1.22 - 2.08) for Child B, and 1.82 (CI95 1.34 - 2.31) for Child C. The results for the mean normalized ETP were 0.87 (CI95 0.84 - 0.90) for Child A patients, 0.78 (CI95 0.68 - 0.88) for Child B, and 0.58 (CI95 0.51 - 0.64) for Child C (p<0.001). While the normalized ETP correlated well with both PT and INR for Child B (P<0.05) and C (p<0.01), there were significant differences within the child A patients where no significant correlation could be identified (Pearson correlation 0.203, R2=0.04, p=0.075. We conclude that the fully automated assay for the determination of the endogenous thrombin potential (ETP) on the BCS® System (both Dade Behring, Marburg, Germany) is a potentially interesting test to measure the coagulation abilities in patients with liver cirrhosis which in the future may supersede the PT/INR in classification systems for hepatic disease.

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