Revised methodology for a blood-clotting response test for identification of warfarin-resistant Norway rats (Rattus norvegicus)

A blood-clotting response test for warfarin resistance in Norway rats ( Rattus norvegicus ) was developed by our laboratory in the late 1970s, based on knowledge of biochemical mechanisms available at that time. That test used warfarin, its the free base which is not watersoluble, and vitamin K₁ 2,3-epoxide as an antidote for warfarin-resistant rats only. We have modified the above test to use a water-soluble salt of vitamin K₃ (1 mg kgg⁻¹ body weight), which is an antidote in a wider range of warfarin-resistant rat strains than vitamin K₁ 2,3- epoxide, and a water-soluble salt of warfarin (5.4 mg kg⁻¹ body weight). Sodium warfarin and vitamin K₃ are administered by oral gavage, and the blood-clotting response measured 24 h later. In a series of validation tests there were no significant differences between results obtained by either method. The new method, however, uses materials which are commercially available, and eliminates the possibility of misclassifying warfarin-resistant rats as susceptible because vitamin K₁ 2,3-epoxide was not an antidote in those strains. We also present evidence that measuring blood-clotting response to feeding on a vitamin K-free diet for 4 days is not a reliable method for determining the genotype of warfarin-resistant rats.