Váradi K, Moritz B, Lang H, Bauer K, Preston E, Peake I, Rivard G E, Keil B, Schwarz H P
Research Laboratories of Immuno AG, Vienna, Austria.
Br J Haematol. 1995 Aug;90(4):884-91. doi: 10.1111/j.1365-2141.1995.tb05210.x.
Resistance to activated protein C (APC) diagnosed on the basis of prolongation of clotting time in an activated partial thromboplastin time (aPTT) assay is now considered a major cause of inherited thrombophilia. The majority of patients with APC resistance carry a factor V molecule with a point mutation at one APC cleavage site (Arg506Gln) which prevents the optimal inactivation of activated factor V by APC. To overcome the limitations of aPTT-based assays in the diagnosis of APC resistance, we have developed a chromogenic assay which is based on the capacity of APC to limit the generation of factor Xa by inactivating factor VIIIa in plasma. The ratio of the factor Xa amidolytic activity in a sample without APC to its factor Xa activity with the addition of APC reflects the response of the plasma coagulation system to APC. The normal range in 44 healthy individuals was 1.62-2.06. APC response ratios as measured by the chromogenic assay correlated with ratios measured by the aPTT assay and were below the normal range in 23/24 individuals with Arg506Gln mutant factor V from three different families with familial thrombosis and from 11 unrelated asymptomatic individuals. In reconstitution experiments, purified factor V corrected the decreased APC response in plasma samples from patients with the Arg506Gln mutation as well as with factor V deficiency, and increased the APC response in normal plasma, whereas the addition of activated factor V had no enhancing effect.
基于活化部分凝血活酶时间（aPTT）检测中凝血时间延长而诊断出的对活化蛋白C（APC）抵抗，现被认为是遗传性易栓症的主要原因。大多数APC抵抗患者携带一种因子V分子，该分子在一个APC裂解位点（Arg506Gln）存在点突变，这阻止了APC对活化因子V的最佳灭活。为克服基于aPTT检测在诊断APC抵抗方面的局限性，我们开发了一种显色检测方法，该方法基于APC通过灭活血浆中的因子VIIIa来限制因子Xa生成的能力。样品在无APC时的因子Xa酰胺水解活性与其加入APC后的因子Xa活性之比反映了血浆凝血系统对APC的反应。44名健康个体的正常范围为1.62 - 2.06。通过显色检测测得的APC反应比值与通过aPTT检测测得的比值相关，并且在来自三个不同家族患有家族性血栓形成的23/24名携带Arg506Gln突变因子V的个体以及11名无关的无症状个体中低于正常范围。在重组实验中，纯化的因子V纠正了来自Arg506Gln突变患者以及因子V缺乏患者血浆样本中降低的APC反应，并增加了正常血浆中的APC反应，而加入活化因子V则没有增强作用。
