alexa A quantitative and humane tail bleeding assay for efficacy evaluation of antihaemophilic factors in haemophilia A mice.


Journal of Blood Disorders & Transfusion

Author(s): Molina ES, Fujita A, Sogayar MC, Demasi MA

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Abstract The tail bleeding model using haemophilic mice has been used as one of the standard assays for efficacy evaluation of novel antihaemophilic therapies at the preclinical level. A number of different configurations and endpoints have been proposed in the literature for this model, hindering interlaboratory comparisons. A particular configuration, known as the tail bleeding survival assay (TBS), adopted by several groups, involves measuring the ability of conscious haemophilic mice to survive exsanguination following tail transection. Major limitations to this configuration include ethical constraints and impaired quantitative determinations. The aim of this study was to standardize and validate a quantitative haemostatic assay for evaluation of antihaemophilic therapies employing an alternative to TBS, which involves a more humane endpoint associated with stable clot formation. Haemophilic mice were treated with vehicle or different doses of two antihaemophilic reference products licensed in Brazil. The haemostatic response was evaluated by our quantitative tail bleeding haemostatic assay (qTBA) over a period of 120 min and then quantified by dose-response modelling. We demonstrate that our qTBA method allows a direct relationship between the number of animals which achieved full haemostatic response and the dosage of both antihaemophilic factors evaluated over 120 min. In addition, the method sensitivity is suitable to demonstrate the conversion from a severe to a moderate haemophilia phenotype. Our proposed qTBA is easy to implement and constitutes an alternative and more ethical endpoint, which could be effectively used as a surrogate to the commonly employed survival endpoint, allowing quantitative haemostatic response evaluation associated with stable clot formation. © 2014 John Wiley & Sons Ltd. This article was published in Haemophilia and referenced in Journal of Blood Disorders & Transfusion

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