Bos R, Laterveer-Vreeswijk G H, Lockwood D, Szewczyk K, Nieuwenhuizen W
TNO Prevention and Health, Gaubius Laboratory, Leiden, The Netherlands.
Thromb Haemost. 1999 Jan;81(1):54-9.
Fibrin formation is a multistep process initiated by thrombin. At first thrombin converts fibrinogen to fibrin molecules which in vivo form soluble complexes with fibrinogen. Soluble fibrin is considered to be an early biochemical marker for intravascular fibrin formation and impending thrombotic events, such as deep venous thrombosis (DVT), pulmonary embolism (PE) and disseminated intravascular coagulopathy (DIC). A new enzyme immunoassay (EIA) was developed on the basis of a monoclonal antibody directed against a fibrin specific neo-epitope located on the gamma-chain of fibrinogen; gamma-(312-324). In addition, it was possible to prepare a lyophilized reference material of thrombin-generated soluble fibrin, that allowed for full antigen recovery after reconstitution with buffer. Assay conditions, e.g. solid phase-Ig concentration and buffer composition, sample and conjugate dilution, and incubation times were optimised. The present assay was found to be specific (no interference of homologous antigens) and reproducible (intra-assay CV 4-8%, interassay CV 4-9%), and therefore highly suited for measuring soluble fibrin levels in a plasma milieu. The median normal value for soluble fibrin was determined in plasma samples obtained from apparently healthy volunteers (n = 81) and found to be 0.040 microg/ml, with a range (10-90 percentiles) of 0.026-0.059 microg/ml. A retrospective study showed that soluble fibrin levels were highly significantly increased in patients with a confirmed diagnosis of DIC (median 1.042 microg FEU/ml, range 0.160-2.319 microg/ml, n = 21, P<0.0001 vs. normal). PE (median 0.527 microg FEU/ml, range 0.084-1.234 microg/ml, n = 29, P<0.0001 vs normal) and DVT (median 0.126 microg FEU/ml, range 0.059-0.878 microg/ml, n = 36, P<0.0001 vs. normal), as determined by the Mann-Whitney U-Test.
纤维蛋白形成是一个由凝血酶启动的多步骤过程。首先，凝血酶将纤维蛋白原转化为纤维蛋白分子，这些纤维蛋白分子在体内与纤维蛋白原形成可溶性复合物。可溶性纤维蛋白被认为是血管内纤维蛋白形成以及即将发生的血栓形成事件（如深静脉血栓形成（DVT）、肺栓塞（PE）和弥散性血管内凝血（DIC））的早期生化标志物。基于针对位于纤维蛋白原γ链上的纤维蛋白特异性新表位（γ-(312 - 324)）的单克隆抗体，开发了一种新的酶免疫测定法（EIA）。此外，还制备了凝血酶生成的可溶性纤维蛋白冻干参考物质，用缓冲液复溶后可实现完全抗原回收。对测定条件，如固相免疫球蛋白浓度、缓冲液组成、样本和结合物稀释度以及孵育时间进行了优化。发现本测定法具有特异性（无同源抗原干扰）和可重复性（批内变异系数4 - 8%，批间变异系数4 - 9%），因此非常适合在血浆环境中测量可溶性纤维蛋白水平。在从明显健康的志愿者（n = 81）获取的血浆样本中测定了可溶性纤维蛋白的正常中位值，发现为0.040微克/毫升，范围（第10 - 90百分位数）为0.026 - 0.059微克/毫升。一项回顾性研究表明，确诊为DIC的患者中可溶性纤维蛋白水平显著升高（中位值1.042微克FEU/毫升，范围0.160 - 2.319微克/毫升，n = 21，与正常相比P<0.0001）。通过曼 - 惠特尼U检验确定，PE患者（中位值0.527微克FEU/毫升，范围0.084 - 1.234微克/毫升，n = 29，与正常相比P<0.0001）和DVT患者（中位值0.126微克FEU/毫升，范围0.059 - 0.878微克/毫升，n = 36，与正常相比P<0.0001）的可溶性纤维蛋白水平也显著升高。
