Ticks are blood-feeding arthropods widely distributed in the world and vectors of several diseases. As haematophagy demands evasion strategies and repeatedly infested hosts develop protective immune responses, we investigated the mechanisms of the Rhipicephalus (Boophilus) microplus saliva anti-haemostatic activity and the possible relationship between the acquired natural anti-tick host resistance and anti-haemostatic action. For this purpose, we studied the effects of R. microplus saliva on different pathways of haemostasis and tested whether repeated infested bovine sera (RIBS) are able to abolish salivary anti-haemostatic activities. R. microplus saliva (i) displays inhibitory activity upon collagen-induced platelet aggregation; (ii) inhibits the induction of endothelial pro-coagulant state; and (iii) reduces thrombogenesis in vivo. RIBS were shown to be able to partially block the delay of coagulation and the anti-thrombotic effect of saliva, and to totally abolish the modulation of endothelium activation. Conversely, RIBS has no effect on the inhibition of platelet aggregation. These results show, for the first time, the neutralization ability of sera from acquired resistance hosts against tick anti-haemostatics. Moreover, this is the first report of a haematophagous parasite able to modulate endothelial cell pro-coagulant state, and addresses the presence of anti-platelet and anti-thrombotic activity in R. microplus saliva.