Antiphospholipid syndrome (APS) is an autoimmune disorder in which vascular thrombosis and recurrent pregnancy loss occur in patients with antiphospholipid antibodies(aPL). Measurements of the beta2-glycoprotein I-dependent anticardiolipin antibody(aCL) and lupus anticoagulant(LAC) are the only laboratory tests available for the diagnosis of APS. Recently, phosphatidylserine-dependent antiprothrombin antibody(aPS/PT) has been detected. aPS/PT was measured by ELISA using the phosphatidylserine-prothrombin complex as an antigen immobilized on ELISA plates in the presence of CaCl2. In our study of 219 patients with APS and autoimmune diseases, the prevalence of aPS/PT-IgG in those with APS was 42.2%, which was significantly higher than that(4.6%) in patients with autoimmune diseases. Furthermore, aPS/PT was closely associated with APS manifestations with an odds ratio (OR) of 2.92 (95% confidence interval (95% CI): 1.33 to approximately 6.40), whereas the OR for aCL was 2.06 (95% CI: 0.91 to approximately 4.66). In addition, aPS/PT-IgG was strongly correlated with the presence of LAC as detected with a diluted Russell viper venom time test (dRVVT) (OR: 38.2, 95% CI: 13.4 to approximately 109.1). The monoclonal antibody (23-1D) of aPS/PT also prolonged the clotting time in LAC tests (aPTT, dRVVT, and kaolin clotting time) in a concentration-dependent manner. In conclusion, aPS/PT is more closely associated with manifestations of APS and LAC, and positive results from an aPS/PT test can mark thrombotic events in APS patients. The determination of aPS/PT in clinical practice, in conjunction with that of other aPL, may improve the likelihood of recognizing APS.