Enzyme-linked immunosorbent assay for the determination of blood coagulation factor XIII A-subunit in plasma and in cell lysates
Enzyme-linked immunosorbent assay for the determination of blood coagulation factor XIII A-subunit in plasma and in cell lysates
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DOI：
10.1016/s0022-1759(01)00479-3
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发表时间：
2001-12-01
影响因子：
2.2
通讯作者：
Muszbek, L
中科院分区：
文献类型：
--
作者：
Katona, É;Ajzner, É;Muszbek, L
建立了一种测定血浆和细胞裂解物中凝血因子XIII亚单位A（FXIII-A）浓度的一步法ELISA新方法。针对FXIII-A上不同表位的单克隆抗体用于测定。捕获抗体在其碳水化合物部分上生物素化，检测抗体用辣根过氧化物酶标记。抗原-抗体反应在链霉亲和素包被的微孔板的孔中进行。与FXIII亚基B（FXIII-B）的复合物形成以及与纤维蛋白原的结合不影响抗体对FXIII-A的可及性。该方法可在2 h内完成，并表现出良好的重现性，回收率和灵敏度。血浆样品在-20 ℃下储存至少6个月后可进行测定。然而，在血小板裂解物的情况下，冷冻和再解冻导致FXIII-A的显著损失。在健康个体和患者的血浆样品中测量的FXIII-A浓度与复合的血浆FXIII（A（2）B（2））的浓度以及与FXIII活性测量的结果良好相关。血小板FXIII-A的参考范围为46-82 fg/血小板。(C)2001 Elsevier Science B. V.保留所有权利。
A new one-step ELISA was developed for the determination of the concentration of blood coagulation factor XIII subunit A (FXIII-A) in plasma and in cell lysates. Monoclonal antibodies directed against different epitopes on FXIII-A were used for the assay. The capture antibody was biotinylated on its carbohydrate moiety and the detection antibody was labelled with horseradish peroxidase. The antigen-antibody reaction was carried out in the well of a streptavidin-coated microplate. Complex formation with FXIII subunit B (FXIII-B) and association to fibrinogen did not influence the accessibility of the antibodies to FXIII-A. The method could be performed within 2 h and demonstrated good reproducibility, recovery and sensitivity. Plasma samples could be assayed after storage at - 20 degreesC for at least 6 months. However, in the case of platelet lysates freezing and rethawing resulted in a significant loss of FXIII-A. FXIII-A concentrations measured in the plasma samples of healthy individuals and patients correlated well with the concentrations of complexed plasma FXIII (A(2)B(2)) and with the results of FXIII activity measurements. A reference range of 46-82 fg/platelet was established for platelet FXIII-A. (C) 2001 Elsevier Science B.V. All rights reserved.