Favaloro E J, Mirochnik O, McDonald D
Department of Haematology, Westmead Hospital, Western Sydney Area Health Service, Australia.
Br J Biomed Sci. 1999;56(1):23-33.
We compare results of factor V DNA analysis with three different clotting-based assays designed to detect activated protein C (APC) resistance (APCR), using samples from 958 patients undergoing assessment for thrombophilia. The original and most commonly used APTT-based procedure (generating an APTT ratio in presence versus absence of APC), showed the least correlation with DNA findings, with a large overlap between normals and heterozygotes. Using this procedure, over 40% of patients with a normal DNA pattern gave APTT ratio results within the heterozygotes' ratio range, and thus is a poor predictor for factor V DNA Leiden mutation (sensitivity 94.3%, specificity 47.0% [APC ratio cut-off: 3.1]; sensitivity 52.1%, specificity 92.9% [APC ratio cut-off: 2.0]). Two commercially available procedures (protein C impedance [PCI] test and protein C pathway [PCP] test), using modified Russell's viper venom time (RVVT) assays, showed less overlap between normals and heterozygotes than did the APTT-based method. Fewer than 10% of normal individuals gave PCI or PCP test ratio results that fell within the respective heterozygotes' ratio range (PCI: sensitivity 95.3%, specificity 96.0%; PCP: sensitivity 97.3%, specificity 82.4% [APC ratio cut-off: 1.6 and 1.9 respectively]). Use of previously described normalisation procedures (patient's APTT ratio over pooled normal plasma [PNP] APTT ratio) showed little improvement in discriminatory power (sensitivity 96.4%, specificity 44.8% [normalised APC ratio cut-off value: 0.97]; sensitivity 58.8%, specificity 90.1% [normalised APC ratio cut-off: 0.68]). Use of factor V-deficient plasma as sample diluent improved discrimination for all assays, but added considerable time and cost to the testing process. Furthermore, use of factor V-deficient plasma dilutions in the APTT-based test (sensitivity 97.1%, specificity 93.8% [APC ratio cut-off: 2.0]) did not substantially improve discrimination compared with either PCI or PCP performed without factor V-deficient plasma. Overall, a combination of RVVT- and APTT-based tests was found to provide excellent discrimination, particularly negative prediction, with respect to the likely factor V DNA result. Of 567 patients co-tested, all factor V DNA-normal patients (n = 299) gave both PCP-RVVT and APCR-APTT (not prediluted with factor V-deficient plasma) test ratio values > or = 2.2. In conclusion, it is important to recognise the limitation of plasma-based assays, in particular the APTT procedure, to discriminate the factor V mutation.
我们使用来自958名接受血栓形成倾向评估患者的样本，将因子V DNA分析结果与三种不同的基于凝血的检测方法进行比较，这些方法旨在检测活化蛋白C（APC）抵抗（APCR）。最初且最常用的基于活化部分凝血活酶时间（APTT）的方法（在有或没有APC的情况下生成APTT比值）与DNA检测结果的相关性最小，正常人和杂合子之间有很大重叠。使用该方法，超过40%DNA模式正常的患者给出的APTT比值结果在杂合子比值范围内，因此它对于因子V DNA莱顿突变是一个较差的预测指标（敏感性94.3%，特异性47.0%[APC比值临界值：3.1]；敏感性52.1%，特异性92.9%[APC比值临界值：2.0]）。两种市售方法（蛋白C阻抗[PCI]试验和蛋白C途径[PCP]试验），使用改良的罗素蝰蛇毒时间（RVVT）检测，与基于APTT的方法相比，正常人和杂合子之间的重叠较少。不到10%的正常个体给出的PCI或PCP试验比值结果落在各自杂合子的比值范围内（PCI：敏感性95.3%，特异性96.0%；PCP：敏感性97.3%，特异性82.4%[APC比值临界值分别为1.6和1.9]）。使用先前描述的标准化程序（患者的APTT比值除以混合正常血浆[PNP]的APTT比值）在鉴别能力上几乎没有改善（敏感性96.4%，特异性44.8%[标准化APC比值临界值：0.97]；敏感性58.8%，特异性90.1%[标准化APC比值临界值：0.68]）。使用因子V缺乏血浆作为样本稀释剂可改善所有检测的鉴别能力，但会给检测过程增加大量时间和成本。此外，在基于APTT的检测中使用因子V缺乏血浆稀释液（敏感性97.1%，特异性93.8%[APC比值临界值：2.0]）与不使用因子V缺乏血浆进行的PCI或PCP相比，在鉴别能力上没有实质性改善。总体而言，发现基于RVVT和APTT的检测方法组合在预测因子V DNA可能结果方面具有出色的鉴别能力，尤其是阴性预测能力。在共同检测的567名患者中，所有因子V DNA正常的患者（n = 299）的PCP - RVVT和APCR - APTT（未用因子V缺乏血浆预稀释）试验比值均≥2.2。总之，认识到基于血浆的检测方法，特别是APTT程序在鉴别因子V突变方面的局限性很重要。
