D'Angelo S V, Gilardoni F, D'Angelo A
Coagulation Service, Scientific Institute IRCCS H.S. Raffaele, Milano, Italy.
Thromb Haemost. 1989 Nov 24;62(3):861-7.
In vivo expression of protein C activity is dependent on the availability of the activated protein C (APC) cofactor protein S. In the clinical laboratory, measurement of protein C anticoagulant activity is mostly performed in modified APTT assays. We have evaluated 13 commercial APTT reagents for their sensitivity to the cofactor effect of protein S by comparing APC-dependent clotting time prolongations in normal plasma and in protein S depleted plasma. In normal plasma, the sensitivities of the APTT reagents to the anticoagulant effect of APC were markedly different and correlated with the sensitivity of reagents to factor V and VIII. Reagents containing soy phosphatides appeared more sensitive than reagents containing phospholipid of animal origin. Analysis of dose-response curves obtained in normal plasma distinguished one group of reagents showing clotting time prolongations linearly related to the APC concentrations, a second group showing a log-linear relationship and a third group showing a log-log relationship. In protein S depleted plasma, sensitivity of APTT reagents to APC was in general proportional to that observed in normal plasma. However, for some reagents dose-response curves were qualitatively different in normal and in protein S depleted plasma. With all the APTT reagents, APC-dependent clotting time prolongations corresponding to 30-80% of APC anticoagulant activity observed in normal plasma, were observed in protein S depleted plasma. At variance, in a modified Xa one-stage clotting assay, negligible clotting time prolongations were observed in protein S depleted plasma, indicating that over 90% of the APC anticoagulant activity was protein S dependent in this assay system. Dilution of a relative insensitive APTT reagent effectively increased its sensitivity to the cofactor effect of protein S, suggesting that different phospholipid content and/or composition might be responsible for the different sensitivity of APTT reagents to protein S. These results question the validity of APTT based assays for the identification of qualitative protein C abnormalities with defective interaction with protein S.
蛋白C活性的体内表达取决于活化蛋白C（APC）辅因子蛋白S的可用性。在临床实验室中，蛋白C抗凝活性的测定大多在改良的活化部分凝血活酶时间（APTT）试验中进行。我们通过比较正常血浆和蛋白S缺乏血浆中APC依赖性凝血时间的延长情况，评估了13种商用APTT试剂对蛋白S辅因子作用的敏感性。在正常血浆中，APTT试剂对APC抗凝作用的敏感性明显不同，且与试剂对因子V和VIII的敏感性相关。含有大豆磷脂的试剂似乎比含有动物源性磷脂的试剂更敏感。对在正常血浆中获得的剂量反应曲线进行分析，区分出一组试剂，其凝血时间延长与APC浓度呈线性关系；第二组呈对数线性关系；第三组呈对数对数关系。在蛋白S缺乏血浆中，APTT试剂对APC的敏感性总体上与在正常血浆中观察到的情况成比例。然而，对于一些试剂，正常血浆和蛋白S缺乏血浆中的剂量反应曲线在质量上有所不同。使用所有APTT试剂时，在蛋白S缺乏血浆中观察到的APC依赖性凝血时间延长相当于正常血浆中观察到的APC抗凝活性的30%-80%。与此不同的是，在改良的Xa一步凝血试验中，在蛋白S缺乏血浆中观察到的凝血时间延长可忽略不计，这表明在该试验系统中，超过90%的APC抗凝活性依赖于蛋白S。稀释相对不敏感的APTT试剂可有效提高其对蛋白S辅因子作用的敏感性，这表明不同的磷脂含量和/或组成可能是APTT试剂对蛋白S敏感性不同的原因。这些结果质疑了基于APTT的试验用于识别与蛋白S相互作用缺陷的定性蛋白C异常的有效性。
