Development of an ELISA for Autoantibodies to Prothrombin Showing their Prevalence in Patients with Lupus Anticoagulants
Development of an ELISA for Autoantibodies to Prothrombin Showing their Prevalence in Patients with Lupus Anticoagulants
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DOI：
10.1055/s-0038-1649891
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发表时间：
1995-10
影响因子：
6.7
通讯作者：
J. Arvieux;L. Darnige;C. Caron;G. Reber;J. Bensa;M. Colomb
中科院分区：
文献类型：
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作者：
J. Arvieux;L. Darnige;C. Caron;G. Reber;J. Bensa;M. Colomb
摘要 一些狼疮抗凝剂 (LA) 已被证明可针对磷脂结合的凝血酶原。在开发 ELISA 来检测患者血清或血浆中的抗凝血酶原自身抗体时，使用固定在普通聚苯乙烯板上的人凝血酶原没有观察到信号或观察到非常低的信号。相比之下，在不添加钙和磷脂的情况下，根据辐射剂量，相同的 LA 阳性样品会与涂覆在 γ 辐射板上的凝血酶原特异性结合。测定的优化需要在缓冲液中添加 0.1% Tween 20。由于液相凝血酶原的竞争性较差，因此通过使用脂质体结合的凝血酶原进行竞争来确定抗体对固定化凝血酶原的特异性。 139 名与多种基础疾病相关的 LA 患者中，有 77 名 (55.4%) 拥有抗凝血酶原抗体（27 种 IgG、35 种 IgM 和 15 种两种同种型），这些抗体要么是分离的，要么更常见的是与抗 (β2 糖蛋白 I (β2GPI) 抗体相关。其中包括 67-71% 的系统性红斑狼疮及相关疾病、原发性抗磷脂抗体综合征患者或药物诱导的 LA（自身免疫组），但只有 19-20% 的患者患有感染或恶性肿瘤（p <0.001），如先前针对抗 β2GPI 抗体所示，IgG2 是与凝血酶原反应的主要 IgG 亚类，因此，患有 LA 的自身免疫患者具有较高的 β2GPI 和凝血酶原抗体发生率，其结合可能同样需要高抗原密度和/或隐秘的暴露。由蛋白质与经辐照（即更多阴离子）聚苯乙烯表面相互作用产生的表位。
Summary Some lupus anticoagulants (LA) have been shown to be directed against phospholipid-bound prothrombin. While developing an ELISA to detect anti-prothrombin autoantibodies in patient serum or plasma, no or very low signal was observed using human prothrombin immobilized on plain polystyrene plates. In contrast, the same LA-positive samples bound specifically to prothrombin coated on γ-irradiated plates, depending on the radiation dose, in the absence of added calcium and phospholipid. Optimization of the assay required the addition of 0.1% Tween 20 to the buffers. Antibody specificity for immobilized prothrombin was ascertained by competition using liposome-bound prothrombin, since fluid-phase prothrombin competed poorly. Seventy-seven of 139 patients (55.4%) with LA related to a variety of underlying diseases possessed anti-prothrombin antibodies (27 IgG, 35 IgM and 15 both isotypes), either isolated or more often associated with anti-(β2 glycoprotein I (β2GPI) antibodies. These included 67-71% of the patients with systemic lupus erythematosus and related disorders, primary antiphospholipid antibody syndrome or drug-induced LA (autoimmune groups), but only 19-20% of those with infection or malignancy (p <0.001). As previously shown for anti-β2GPI antibodies, IgG2 was the predominant IgG subclass reactive with prothrombin. Thus, autoimmune patients with LA have a high incidence of antibodies to β2GPI and prothrombin, the binding of which could similarly require high antigen density and/or exposure of cryptic epitopes resulting from protein interaction with an irradiated (i. e. more anionic) polystyrene surface.