The thrombin-inactivating α2-globulin, antithrombin III (At-III), was measured in plasma and urine with a chromogenic tripeptide substrate, using a centrifugal analyzer. Plasma At-III was subnormal in liver disease. In acute myocardial infarction a mean At-III of approximately 120% of normal was found. At-III in persons who developed deep-vein thrombosis decreased significantly (p < 0.05) between the day of admission and the 3rd day after admission to hospital. Urine from normals did not contain detectable At-III. With increasing urinary albumin increasing concentration of At-III was found