Antibodies directed against protein S (anti-ProtS) may be involved in the development of thrombosis in patients with the antiphospholipid syndrome. We assessed the prevalence and clinical significance of anti-ProtS and evaluated their immunological characteristics in 184 patients with SLE and 99 healthy donors. All patients were tested for IgG anti-ProtS by an in-house ELISA. Plasma levels and functional activity of protein S were also tested. Anti-ProtS were found in 57 patients (31%) and 4 healthy controls (4%). Patients with thrombosis had anti-ProtS more frequently than controls (29% vs 4%, OR 9.5 [95% CI 3.07-29.3], p<0.0001). Anti-ProtS were more frequent in patients with venous thrombosis and in those with arterial thrombosis, than in controls (41% vs. 4%, OR 16.5 [95% CI 5-54], p<0.0001 and 23% vs. 4%, OR 7 [95%CI 2.1-23.5], p=0.0008, respectively). Patients with prematurity, preeclampsia and intrauterine growth restriction had anti-ProtS more frequently than the control group (36%, 47% and 44% vs. 4%; OR 13.6 [95% CI 2.8-66], p=0.003, OR 21 [95% CI 5-86], p<0.0001 and OR 19 [95% CI 4-99], p=0.0014, respectively). Plasma levels of free protein S were not statistically different between patients with and without anti-ProtS and controls (77.9% [20.7-100] vs. 83.7% [52.7-100] vs. 89% [62-101], respectively). Free protein S functional activity was no different between subgroups (105% [48-230] in anti-ProtS positive vs. 123% [95-283] in anti-ProtS negative vs. 136% [60-174] in controls). Anti-ProtS are frequent in SLE patients with thrombosis and pregnancy morbidity. These antibodies do not interfere with free protein S in plasma since its level and/or functional activity are not impaired.