Zherdeva V V, Chudinov A V, Savitskiĭ A P
Bakh Institute of Biochemistry Russian Academy of Sciences, 119071 Moscow, Leninsky pr., 33.
Biomed Khim. 2003 Jan-Feb;49(1):70-9.
The determination of thyroxin (T4) is a basic confirmative test for congenital hypothyroidism. 70% cases of this period inborn disease are taking asymptomatic course. We developed immunofluorescent T4-assay in dried blood spots with anti-T4 monoclonal antibody and europium chelate (dianhydride of diethylentriaminipentaacetic acid (DA-DTPA)). This method requires DELFIA Plate Fluorometer 1232 (Wallac, Finland) or its sub modifications. Panel of monoclonal antibodies for T4 has been obtained. Type/subtype (IgG2b, IgG1), affinity constants (10(7)-10(8) M-1), cross reactivity to homologous structures (0-2%) were determined. Stable clones with high affinity and viability were selected for the development of the assay. Conjugates of thyroxin and europium chelate were synthesized. Inclusion of sodium salicylate (2 mg/ml) and EDTA (2 mM) into the buffer reduced the nonspecific signal. Limit for T4 detection (T4 standarts) was 10 nM with not more than 15% variation coefficient. Accuracy was estimated by Bio-Rad Lipochek Immunoassay Plus Control Kit. Obtained results were within control confidence interval.
甲状腺素（T4）测定是先天性甲状腺功能减退症的一项基本确诊试验。该先天性疾病在此阶段70%的病例呈无症状病程。我们利用抗T4单克隆抗体和铕螯合物（二乙三胺五乙酸二酐（DA - DTPA））开发了干血斑免疫荧光T4测定法。此方法需要1232型DELFIA板式荧光计（芬兰Wallac公司）或其衍生型号。已获得用于T4的单克隆抗体组。测定了其类型/亚型（IgG2b、IgG1）、亲和常数（10⁷ - 10⁸ M⁻¹）以及对同源结构的交叉反应性（0 - 2%）。选择具有高亲和力和活力的稳定克隆用于该测定法的开发。合成了甲状腺素与铕螯合物的结合物。在缓冲液中加入水杨酸钠（2 mg/ml）和乙二胺四乙酸（EDTA，2 mM）可降低非特异性信号。T4检测限（T4标准品）为10 nM，变异系数不超过15%。通过伯乐Lipochek免疫测定增强型对照试剂盒评估准确性。所得结果在对照置信区间内。
