Haemostasis Research Unit, Haematology Department, University College London and University College London Hospitals, 1st Floor, 51 Chenies Mews, London, WC1E 6HX, UK.
Thromb Haemost. 2013 Mar;109(3):488-96. doi: 10.1160/TH12-08-0565. Epub 2013 Jan 10.
ADAMTS13 activity assays are sometimes useful in confirming the clinical diagnosis or to distinguish different thrombotic microangiopathies (TMA). We investigated the commonly used clinical assays for ADAMTS13 activity. 159 samples from normal subjects or acquired TMA patients were studied in collagen binding (CBA), Fret and chromogenic peptide substrate assays. Frozen aliquots of pooled normal plasma gave similar values by CBA, Fret-VWF73 peptide, Fret-VWF86 and chromogenic VWF73 ELISA (chr-VWF73). Two lyophilised commercial calibrants gave lower ADAMTS13 activity by CBA than peptide substrate assays. The addition of solid HEPES to normal plasma caused a significant fall in CBA, but not Fret-VWF73 activity and might partly explain the differences, since lyophilised plasmas are often HEPES buffered. Normal plasmas showed good agreement between CBA and Fret assays, although chr-VWF73 gave slightly higher values. In acquired TMA, there was reasonable agreement between assays for samples with <11% ADAMTS13 activity (83% of samples showed agreement between CBA, Fret-VWF73 and chr-VWF73), but samples with moderate deficiency frequently showed lower CBA levels (only 41-52% agreement). However, there were also some discrepancies among the peptide substrate assays, with Fret-VWF86 sometimes giving slightly higher values than the VWF73 substrate assays. An International reference plasma might improve standardisation, but is not the only problem. It is unclear which assay has greatest clinical utility, this may depend on the nature of the sample. If the activity does not match the clinical picture, an alternative method should be performed. Where therapeutic monitoring is required, the same activity assay should be used throughout.
ADAMTS13 活性测定有时有助于确认临床诊断或区分不同的血栓性微血管病（TMA）。我们研究了常用的 ADAMTS13 活性临床测定方法。对 159 例来自正常受试者或获得性 TMA 患者的样本，使用胶原结合（CBA）、Fret 和显色肽底物测定法进行了研究。通过 CBA、Fret-VWF73 肽、Fret-VWF86 和显色 VWF73 ELISA（chr-VWF73），对来自正常混合血浆的冷冻等分试样进行了研究，结果相似。两种冻干的商业校准品通过 CBA 测定的 ADAMTS13 活性低于肽底物测定法。向正常血浆中添加固体 HEPES 会导致 CBA 活性显著下降，但不会影响 Fret-VWF73 活性，这可能部分解释了差异，因为冻干血浆通常是 HEPES 缓冲的。正常血浆的 CBA 和 Fret 测定之间具有良好的一致性，尽管 chr-VWF73 给出的数值略高。在获得性 TMA 中，对于 ADAMTS13 活性<11%的样本（83%的样本在 CBA、Fret-VWF73 和 chr-VWF73 之间具有一致性），各测定法之间具有合理的一致性，但中度缺乏的样本通常显示较低的 CBA 水平（仅 41-52%的一致性）。然而，肽底物测定法之间也存在一些差异，Fret-VWF86 有时比 VWF73 底物测定法给出略高的值。国际参考血浆可能会改善标准化，但这并不是唯一的问题。尚不清楚哪种测定法具有最大的临床实用性，这可能取决于样本的性质。如果活性与临床情况不匹配，应进行替代方法。如果需要治疗监测，则应在整个过程中使用相同的活性测定法。
