The objective of the present study was to evaluate the influence of thrombophilia risk factors on variables of a chromogenic thrombin generation assay (ETP) in a setting with acute deep venous thrombosis (DVT) and non-DVT patients. In 152 outpatients suspected for DVT, the results of thrombophilia investigations were known. In all patients, thrombin generation parameters were determined and related to the presence of thrombophilia risk factors. The thrombin generation results were divided in quartiles to calculate the odds ratio, as a measure of the association with thrombophilia risk factors, corrected for age and sex. The groups with and without DVT were analysed independently. From the 152 patients, there were 108 patients with and 44 without DVT. In the DVT-positive group, lag time was significantly prolonged (21.9 versus 20.3 s; P=0.021) in patients with known thrombophilia risk factors (n=48). The odds ratio for thrombophilia risk factors was 3.7 (95% CI 1.1-12) uncorrected and 4.3 (95% CI 1.2-16) corrected for age and sex, both P values less than 0.05. No differences in ETP parameters were observed in patients without DVT with known thrombophilia risk factors (n=32). The relationship of thrombophilia risk factors with the occurrence of a prolonged lag time in the acute phase of DVT is a new finding. Increased coagulation activation may be the consequence of the thrombophilia risk factors with this observation. To unravel whether the test expresses increased consumption or increased release of anticoagulant factors may open new ways to develop diagnostic methods to add to further refinement of exclusion algorithms for DVT.