Name: De Nijmegen Hemostasis-assay: gelijktijdige fluorogene meting van trombine- en plasminegeneratie in één put (Video) | JoVE Journal
Uploaded: 2026-02-27T13:15:00.000+00:00
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Description: Leer de Nijmegen Hemostasis-assay voor gelijktijdige fluorogene meting van trombine- en plasminegeneratie in plasma. Deze methode biedt een geïntegreerde beoordeling van hemostase en fibrinolyse.

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<li>Tripodi, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Thrombin+generation+assay+and+its+application+in+the+clinical+laboratory%5BTitle%5D)+AND+%22Clin+Chem%22%5BJournal%5D)">Thrombin generation assay and its application in the clinical laboratory.</a> Clin Chem. 62 (5), 699-707 (2016).</li>
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<li>van Geffen, M., van Heerde, W. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Global+haemostasis+assays%2C+from+bench+to+bedside%5BTitle%5D)+AND+%22Thromb+Res%22%5BJournal%5D)">Global haemostasis assays, from bench to bedside.</a> Thromb Res. 129 (6), 681-687 (2012).</li>
<li>Luddington, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Thrombelastography%2Fthromboelastometry%5BTitle%5D)+AND+%22Clin+Lab+Haematol%22%5BJournal%5D)">Thrombelastography/thromboelastometry.</a> Clin Lab Haematol. 27 (2), 81-90 (2005).</li>
<li>Hartmann, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Viscoelastic+testing%3Aan+illustrated+review+of+technology+and+clinical+applications%5BTitle%5D)+AND+%22Res+Pract+Thromb+Haemost%22%5BJournal%5D)">Viscoelastic testing:an illustrated review of technology and clinical applications.</a> Res Pract Thromb Haemost. 7 (1), 100031(2023).</li>
<li>Schoeman, R. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Flow+chamber+and+microfluidic+approaches+for+measuring+thrombus+formation+in+genetic+bleeding+disorders%5BTitle%5D)+AND+%22Platelets%22%5BJournal%5D)">Flow chamber and microfluidic approaches for measuring thrombus formation in genetic bleeding disorders.</a> Platelets. 28 (5), 463-471 (2017).</li>
<li>Bavinck, A. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Viscoelastic+testing+in+patients+with+very+rare+bleeding+disorders+of+secondary+hemostasis%3Aa+scoping+review%5BTitle%5D)+AND+%22Thromb+Res%22%5BJournal%5D)">Viscoelastic testing in patients with very rare bleeding disorders of secondary hemostasis:a scoping review.</a> Thromb Res. 2025, 109449(2025).</li>
<li>Mansouritorghabeh, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+utility+of+total+thrombus-formation+analysis+system+%28T-TAS%29+in+the+thrombosis+and+hemostasis+field%3Aa+scoping+review%5BTitle%5D)+AND+%22Int+J+Lab+Hematol%22%5BJournal%5D)">The utility of total thrombus-formation analysis system (T-TAS) in the thrombosis and hemostasis field:a scoping review.</a> Int J Lab Hematol. 47 (2), 201-211 (2025).</li>
<li>Lecchi, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Flow-chamber+device+%28T-TAS%29+to+diagnose+patients+suspected+of+platelet+function+defects%5BTitle%5D)+AND+%22Blood+Transfus%22%5BJournal%5D)">Flow-chamber device (T-TAS) to diagnose patients suspected of platelet function defects.</a> Blood Transfus. 22 (1), 55-64 (2024).</li>
<li>Zekavat, O. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Comparison+of+thrombin+generation+assay+with+conventional+coagulation+tests+in+evaluation+of+bleeding+risk+in+patients+with+rare+bleeding+disorders%5BTitle%5D)+AND+%22Clin+Appl+Thromb+Hemost%22%5BJournal%5D)">Comparison of thrombin generation assay with conventional coagulation tests in evaluation of bleeding risk in patients with rare bleeding disorders.</a> Clin Appl Thromb Hemost. 20 (6), 637-644 (2014).</li>
<li>Brummel-Ziedins, K. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Thrombin+generation+and+bleeding+in+haemophilia+A%5BTitle%5D)+AND+%22Haemophilia%22%5BJournal%5D)">Thrombin generation and bleeding in haemophilia A.</a> Haemophilia. 15 (5), 1118-1125 (2009).</li>
<li>Al Dieri, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Thrombin+generation%3Awhat+have+we+learned%5BTitle%5D)+AND+%22Blood+Rev%22%5BJournal%5D)">Thrombin generation:what have we learned.</a> Blood Rev. 26 (5), 197-203 (2012).</li>
<li>Binder, N. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Clinical+use+of+thrombin+generation+assays%5BTitle%5D)+AND+%22J+Thromb+Haemost%22%5BJournal%5D)">Clinical use of thrombin generation assays.</a> J Thromb Haemost. 19 (12), 2918-2929 (2021).</li>
<li>Verhagen, M. J. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((In+patients+with+hemophilia%2C+a+decreased+thrombin+generation+profile+is+associated+with+a+severe+bleeding+phenotype%5BTitle%5D)+AND+%22Res+Pract+Thromb+Haemost%22%5BJournal%5D)">In patients with hemophilia, a decreased thrombin generation profile is associated with a severe bleeding phenotype.</a> Res Pract Thromb Haemost. 7 (2), 100062(2023).</li>
<li>Haisma, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Enhanced+thrombin+and+plasmin+generation+profiles+in+alpha-2-antiplasmin-deficient+patients%3Adata+from+the+Rare+Bleeding+Disorders+in+the+Netherlands+study%5BTitle%5D)+AND+%22Res+Pract+Thromb+Haemost%22%5BJournal%5D)">Enhanced thrombin and plasmin generation profiles in alpha-2-antiplasmin-deficient patients:data from the Rare Bleeding Disorders in the Netherlands study.</a> Res Pract Thromb Haemost. 8 (7), 102604(2024).</li>
<li>Haisma, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Comparative+analysis+of+thrombin+generation+platforms+for+patients+with+coagulation+factor+deficiencies%3Aa+comprehensive+assessment%5BTitle%5D)+AND+%22Thromb+Res%22%5BJournal%5D)">Comparative analysis of thrombin generation platforms for patients with coagulation factor deficiencies:a comprehensive assessment.</a> Thromb Res. 240, 109045(2024).</li>
<li>van Geffen, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Retrospective+evaluation+of+bleeding+tendency+and+simultaneous+thrombin+and+plasmin+generation+in+patients+with+rare+bleeding+disorders%5BTitle%5D)+AND+%22Haemophilia%22%5BJournal%5D)">Retrospective evaluation of bleeding tendency and simultaneous thrombin and plasmin generation in patients with rare bleeding disorders.</a> Haemophilia. 18 (4), 630-638 (2012).</li>
<li>Haisma, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((genotype%2C+and+laboratory+assessment+of+congenital+fibrinogen+disorders%3Adata+from+the+Rare+Bleeding+Disorders+in+the+Netherlands+study%5BTitle%5D)+AND+%22Thromb+Res%22%5BJournal%5D)">genotype, and laboratory assessment of congenital fibrinogen disorders:data from the Rare Bleeding Disorders in the Netherlands study.</a> Thromb Res. 249, 109317(2025).</li>
<li>Bouma, B. N., Meijers, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Thrombin-activatable+fibrinolysis+inhibitor+%28TAFI%2C+plasma+procarboxypeptidase+B%2C+procarboxypeptidase+R%2C+procarboxypeptidase+U%29%5BTitle%5D)+AND+%22J+Thromb+Haemost%22%5BJournal%5D)">Thrombin-activatable fibrinolysis inhibitor (TAFI, plasma procarboxypeptidase B, procarboxypeptidase R, procarboxypeptidase U).</a> J Thromb Haemost. 1 (7), 1566-1574 (2003).</li>
<li>Marx, P. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Thrombin-activatable+fibrinolysis+inhibitor%5BTitle%5D)+AND+%22Curr+Med+Chem%22%5BJournal%5D)">Thrombin-activatable fibrinolysis inhibitor.</a> Curr Med Chem. 11 (17), 2335-2348 (2004).</li>
<li>Rijken, D. C., Lijnen, H. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((New+insights+into+the+molecular+mechanisms+of+the+fibrinolytic+system%5BTitle%5D)+AND+%22J+Thromb+Haemost%22%5BJournal%5D)">New insights into the molecular mechanisms of the fibrinolytic system.</a> J Thromb Haemost. 7 (1), 4-13 (2009).</li>
<li>Depasse, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Thrombin+generation+assays+are+versatile+tools+in+blood+coagulation+analysis%3Aa+review+of+technical+features%2C+and+applications+from+research+to+laboratory+routine%5BTitle%5D)+AND+%22J+Thromb+Haemost%22%5BJournal%5D)">Thrombin generation assays are versatile tools in blood coagulation analysis:a review of technical features, and applications from research to laboratory routine.</a> J Thromb Haemost. 19 (12), 2907-2916 (2021).</li>
<li>Subcommittee on Control of Anticoagulation of the SSC of the ISTH. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Towards+a+recommendation+for+the+standardization+of+the+measurement+of+platelet-dependent+thrombin+generation%5BTitle%5D)+AND+%22J+Thromb+Haemost%22%5BJournal%5D)">Towards a recommendation for the standardization of the measurement of platelet-dependent thrombin generation.</a> J Thromb Haemost. 9 (9), 1859-1861 (2011).</li>
<li>Duarte, R. C. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Standardization+and+evaluation+of+the+performance+of+the+thrombin+generation+test+under+hypo-+and+hypercoagulability+conditions%5BTitle%5D)+AND+%22Hematol+Transfus+Cell+Ther%22%5BJournal%5D)">Standardization and evaluation of the performance of the thrombin generation test under hypo- and hypercoagulability conditions.</a> Hematol Transfus Cell Ther. 41 (3), 244-252 (2019).</li>
<li>Dargaud, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Effect+of+standardization+and+normalization+on+imprecision+of+calibrated+automated+thrombography%3Aan+international+multicentre+study%5BTitle%5D)+AND+%22Br+J+Haematol%22%5BJournal%5D)">Effect of standardization and normalization on imprecision of calibrated automated thrombography:an international multicentre study.</a> Br J Haematol. 139 (2), 303-309 (2007).</li>
<li>Ninivaggi, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Recommendations+for+the+measurement+of+thrombin+generation%3Acommunication+from+the+ISTH+SSC+Subcommittee+on+Lupus+Anticoagulant%2FAntiphospholipid+Antibodies%5BTitle%5D)+AND+%22J+Thromb+Haemost%22%5BJournal%5D)">Recommendations for the measurement of thrombin generation:communication from the ISTH SSC Subcommittee on Lupus Anticoagulant/Antiphospholipid Antibodies.</a> J Thromb Haemost. 19 (5), 1372-1378 (2021).</li>
<li>Duchemin, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Influence+of+coagulation+factors+and+tissue+factor+concentration+on+the+thrombin+generation+test+in+plasma%5BTitle%5D)+AND+%22Thromb+Haemost%22%5BJournal%5D)">Influence of coagulation factors and tissue factor concentration on the thrombin generation test in plasma.</a> Thromb Haemost. 99 (4), 767-773 (2008).</li>
<li>Miszta, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+high-fat+diet+delays+plasmin+generation+in+a+thrombomodulin-dependent+manner+in+mice%5BTitle%5D)+AND+%22Blood%22%5BJournal%5D)">A high-fat diet delays plasmin generation in a thrombomodulin-dependent manner in mice.</a> Blood. 135 (19), 1704-1717 (2020).</li>
</ol>

<table><tbody><tr><td>Materiaal</td><td></td><td></td><td></td></tr><tr><td>Calciumchloridedihydraat (CaCl2.2H2O)</td><td>Merck</td><td>1.02382</td><td></td></tr><tr><td>Cephaline (C.K. Prest 5 APTT reagent kit)</td><td>Stago</td><td>00847</td><td>Stabiel gedurende 6 maanden bij -20 &deg;C.&nbsp;</td></tr><tr><td>Gedestilleerd water</td><td></td><td></td><td>Noteer de datum van opening. Gebruik binnen 1 dag na eerste opening.</td></tr><tr><td>Dimethylsulfoxide (DMSO)</td><td>Merck</td><td>1.02950.0500</td><td></td></tr><tr><td>Humaan alfa-trombine</td><td>Enzyme Research Laboratories</td><td>HT 1002a</td><td>De doos bevat een bijsluisbrief met een certificaat van analyse. Het volume buffer dat moet worden toegevoegd, is afhankelijk van de opgegeven concentratie en moet worden aangepast zodat 1 &micro;L toegevoegde humane trombine in 120 &micro;L reagens een eindconcentratie van 76 nmol/L oplevert.</td></tr><tr><td>Humaan plasmine</td><td>Enzyme Research Laboratories</td><td>HPlas</td><td>De doos bevat een bijsluisbrief met een certificaat van analyse. Het volume buffer dat moet worden toegevoegd, is afhankelijk van de opgegeven concentratie en moet worden aangepast zodat 1 &micro;L toegevoegde humane plasmine in 120 &micro;L reagens resulteert in een eindconcentratie van 200 nmol/L.</td></tr><tr><td>Normaal gepoold plasma</td><td>In-house bereiding</td><td>Niet van toepassing</td><td>Stabiel gedurende 3 uur na ontdooien als het op ijs wordt bewaard.</td></tr><tr><td>Patiëntenplasma</td><td>Niet van toepassing</td><td>Niet van toepassing</td><td></td></tr><tr><td>Plasmine substraat (CBZ-L-fenylalanine-L-arginine amide)-rodamine-morfolino ureum</td><td>Symeres</td><td></td><td>Stabiel gedurende 6 maanden bij -80 &deg;C.&nbsp;</td></tr><tr><td>Recombinant humaan weefselfactor (TF): Innovin</td><td>Dade Behring</td><td>B4212</td><td>Stabiel gedurende 6 maanden bij -80 &deg;C.&nbsp;</td></tr><tr><td>Recombinant weefsel-type plasminogenaar activator (tPA): Alteplase/Actilyse</td><td>Boehringer Ingelheim</td><td></td><td>Stabiel gedurende 12 maanden bij -80 &deg;C.&nbsp;</td></tr><tr><td>Tris(hydroxylmethyl)aminomethaan (H2NC(CH2OH)3)</td><td>Merck</td><td>1.08382.2500</td><td></td></tr><tr><td>Tris-NaCl buffer, pH 7,4 (TBS)</td><td>In-house bereiding</td><td>Niet van toepassing</td><td></td></tr><tr><td>Tris-NaCl-CaCl2 buffer, pH 7,4 (TBS-Ca)</td><td>In-house bereiding</td><td>Niet van toepassing</td><td></td></tr><tr><td>Trombine substraat Bz-bèta-Ala-Gly-Arg-7-amino-4-methylcoumarine</td><td>Symeres</td><td></td><td>Stabiel gedurende 6 maanden bij -80 &deg;C.&nbsp;</td></tr><tr><td>Apparatuur</td><td></td><td></td><td></td></tr><tr><td>Fluostar Optima fluorimeter / microplaatlezer met temperatuurregeling en reagentinjectors</td><td>BMG Labtech</td><td></td><td></td></tr><tr><td>96-wells microplaat, polystyreen, vlakke bodem, zwart, medium bindingsoppervlak (Microlon equivalent)</td><td>Greiner BIO-ONE</td><td>655076</td><td></td></tr><tr><td>Analytische weegschaal (bovenlader)</td><td></td><td></td><td></td></tr><tr><td>Blauwe pipettetips (voor 100-1000 &micro;L)</td><td></td><td></td><td></td></tr><tr><td>Citraat Bloedverzamelbuisjes 3mL (3,2% Citraat)</td><td>BD Vacutainer</td><td></td><td></td></tr><tr><td>IJsemmer</td><td></td><td></td><td></td></tr><tr><td>IJsmachine</td><td></td><td></td><td></td></tr><tr><td>Incubator / microplaat shaker</td><td></td><td></td><td></td></tr><tr><td>Magneetroerder</td><td></td><td></td><td></td></tr><tr><td>Microcentrifugebuisjesrek</td><td></td><td></td><td></td></tr><tr><td>Microcentrifugebuisjes, afsluitbaar</td><td></td><td></td><td></td></tr><tr><td>pH-meter</td><td></td><td></td><td></td></tr><tr><td>Pipettips 0,1-10 &micro;L, niet-steriel</td><td></td><td></td><td></td></tr><tr><td>Pipetten (2, 10, 20, 100 en 1000 &micro;L)</td><td></td><td></td><td></td></tr><tr><td>Schroevendraaier (gebruikt als sonde)</td><td></td><td></td><td></td></tr><tr><td>Buismixer</td><td></td><td></td><td></td></tr><tr><td>Waterbad</td><td></td><td></td><td></td></tr><tr><td>Gele pipettetips (voor 20-200 &micro;L)</td><td></td><td></td><td></td></tr></tbody></table>

de nijmegen hemostasis-assay: gelijktijdige fluorogene meting van trombine- en plasminegeneratie in één put

The Nijmegen Hemostasis Assay: Simultaneous Fluorogenic Measurement of Thrombin and Plasmin Generation in a Single Well

De Nijmegen Hemostasis-assay: gelijktijdige fluorogene meting van trombine- en plasminegeneratie in één put

Thrombin and plasmin both play a central role in hemostasis, regulating coagulation and fibrinolysis, respectively. Conventional hemostatic assays often ...

the-nijmegen-hemostasis-assay-simultaneous-fluorogenic-measurement

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Medicine

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Deze studie beschrijft een nieuwe microplaat test die FV stollingsactiviteit tijdens fibrine stolselvorming in menselijk plasma die niet eerder gerapporteerd. De methode gebruikt een kinetische microplaatlezer de verandering in absorptie bij 405 nm meet continu in fibrine stolselvorming in menselijk plasma.

Meting van Factor V activiteit in humaan plasma met een Microplate coagulatietest

Demonstratie van een snelle kwantitatieve analyse van de remming van bloedstolling door de laag moleculair gewicht heparine, enoxaparine. De bijdrage van enoxaparine wordt beoordeeld door het verwijderen van de invloed door digestie met heparinase. Een vollediger beschrijving van de assay wordt beschreven in onze gepubliceerde papier.

Snelle Point-of-Care test van Enoxaparine anticoagulans werkzaamheid in volbloed

This video demonstrates the use of a UV-visible spectrophotometer for monitoring insoluble fibrin clot formation as a function of the change in the solution turbidity. The UV-visible spectrophotometer measures the solution’s absorbance over time, which represents the turbidity change due to the insoluble fibrin clot formation.

UV-Visible Spectroscopy for Monitoring Fibrin Clot Formation

This video demonstrates an assay to study the plasma membrane resealing efficiency in mammalian cells. The cells are exposed to a bacterial pore-forming toxin, which forms pores in the cell membrane. In the presence of extracellular calcium ions, the pore causes calcium influx, inducing membrane-resealing events. The resealing efficiency is assessed using DNA-binding propidium iodide dye, which only enters cells without a resealed membrane.

An In Vitro Fluorescence-Based Assay to Measure Plasma Membrane Resealing Efficiency

Dit artikel beschrijft een experimenteel model van hemostase die gelijktijdig meet bloedplaatjesfunctie en coagulatie. Bloedplaatjes en fibrine fluorescentie wordt gemeten in real-time en bloedplaatjesadhesie rate, coagulatie snelheid, en het begin van coagulatie bepaald. Het model wordt gebruikt voor bloedplaatjes stollingsbevorderende eigenschappen bepalen onder stroom concentraten voor transfusie.

Een microfluïdische Flow Kamer Model voor trombocytentransfusie en hemostase Maatregelen afzetting van trombocyten en fibrine Formation in real-time

Fibrin is verantwoordelijk voor stolselvorming tijdens hemostase en trombose. Troebelheidstesten en tromboelastograhy (TEG) kunnen worden gebruikt als synergetische instrumenten die complementaire beoordeling van een stolsel bieden. Deze twee technieken samen kunnen meer inzicht geven in hoe stollingscondities de vorming van fibrinstolsel beïnvloeden.

Gebruikmaken van troebelheid en trombo-lastografie voor complementaire clot karakterisering

In-vitro trombolyse-assays hebben vaak moeite om in-vivo-omstandigheden na te bootsen, of het nu gaat om de modeltrombus die wordt verteerd of in de omgeving waarin trombolyse plaatsvindt. Hierin onderzoeken we hoe de koppeling van de Chandler-lus en Real-Time Fluorometric Flowing Fibrinolysis Assay (RT-FluFF) wordt gebruikt voor high-fidelity, ex-vivo, stolsellysisbewaking.

Fibrinolyse volgen van Chandler Loop-gevormde volbloedstolsels onder afschuifstroom in een in-vitro trombolysemodel

De bloedplaatjesfunctie onder stroom kan worden beoordeeld en gesimuleerde hemostatische reanimatie kan worden gemodelleerd met behulp van een microfluïdisch apparaat, dat toepassingen heeft in de trauma- en transfusiegeneeskunde.

Microfluïdica bij het beoordelen van de bloedplaatjesfunctie

Het huidige protocol biedt een uitgebreide reeks procedures die nodig zijn om procoagulerende bloedplaatjes te analyseren, die overlappende kenmerken van necrose, apoptose en activering van bloedplaatjes vertonen.

Uitgebreide analyse van procoagulerende bloedplaatjes die kenmerken van necrose, apoptose en activering van bloedplaatjes vertonen

Dit werk beschrijft een microfluissue-assay die schuifspanning gebruikt om de vorming van tromben in het bloed te triggeren en karakteriseert de trombus aan de hand van meerdere dimensies van uitlezen. De test kan de protrombische neiging van bloedmonsters meten en is daardoor nuttig voor ziektediagnose, geneesmiddelenontdekking, evenals basismechanistische studies gerelateerd aan trombose.

Thrombus Profiling Assay: Een op microfludiica gebaseerd platform voor het volledig karakteriseren van biomechanische trombogenese

Leer de Nijmegen Hemostasis-assay voor gelijktijdige fluorogene meting van trombine- en plasminegeneratie in plasma. Deze methode biedt een geïntegreerde beoordeling van hemostase en fibrinolyse.

De Nijmegen Hemostasis-assay: gelijktijdige fluorogene meting van trombine- en plasminegeneratie in één put

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