Antithrombin (AT), a serine protease inhibitor, is involved in inhibiting thrombin and various other coagulation proteases in addition to exerting anti-inflammatory effects. AT deficiency was first described in a family member with venous thromboembolism. The disorder is categorized into two types: type I (qualitative defect) and type II (quantitative defect). The functional assay is the primary test for evaluating AT deficiency, while the antigenic assay can differentiate between types I and II. Although genetic analysis is not routinely performed in a clinical setting, it can be valuable for prenatal diagnosis in cases of homozygous or compound heterozygous AT deficiency. Recent findings indicate that two novel AT variants affecting the N-glycosylation of Asn224 and leading to severe thrombophilia are not detected by standard functional assays. This emphasizes the need for the development of new functional assays and advanced genetic testing to identify these atypical cases of AT deficiency.