The antiphospholipid syndrome (APS) is a thrombotic disorder leading to spontaneous abortions, venous thromboses, myocardial infarctions and strokes. Although the syndrome is associated with characteristic autoantibodies, these tests have poor predictive value for thrombosis. The aim of the study was to determine whether the combined presence of two types of antiphospholipid antibodies can be associated with a high-risk subset of thrombosis-prone patients. One hundred and thirty-four sera from a lupus clinic were tested for antibodies to beta2-glycoprotein I (beta2GPI), protein S and prothrombin. In a group of 29 patients for whom plasma was available, free (functional) protein S levels were also measured. Autoantibodies to beta2GPI and protein S are associated with each other. Dual reactivity to beta2GPI and protein S correlates with increased history of thrombotic events (69% of doubly reactive patients) when compared to either type of autoantibody alone (37% of patients with only anti-beta2GPI and 38% with only anti-protein S, P=0.04 and P=0.01, respectively) or neither reactivity (37%). Among 29 patients tested for free (functional, anticoagulant) protein S levels, the lowest levels were found in patients with antibodies to beta2GPI and/or protein S, and all four patients with a history of thrombosis had below-normal free protein S levels. These associations were not found with antiprothrombin antibodies. In conclusion dual autoantibodies to beta2GPI and protein S are associated with increased history of thrombosis in the antiphospholipid syndrome.