Hereditary antithrombin deficiency is a hypercoagulable state associated with an increased risk for venous thrombosis. The recommended initial test for antithrombin is an activity (functional) assay. The advantages and disadvantages of the various testing options are presented. The causes of acquired antithrombin deficiency are much more common than hereditary deficiency. Therefore, this article describes the appropriate steps to take when antithrombin activity is low, in order to confirm or exclude a hereditary deficiency. The causes of falsely normal results are also described, including direct thrombin inhibitors. Am. J. Hematol. 85:947–950, 2010. © 2010 Wiley-Liss, Inc.