An enzyme immunoassay system with a monoclonal antibody for the determination of 11-deoxycortisol.
An enzyme immunoassay system with a monoclonal antibody for the determination of 11-deoxycortisol.
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带有单克隆抗体的酶免疫分析系统，用于测定 11-脱氧皮质醇。
DOI：
10.1248/cpb.35.1497
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发表时间：
1985
影响因子：
1.7
通讯作者：
T. Nambara
中科院分区：
文献类型：
--
作者：
H. Hosoda;S. Tamura;R. Tsukamoto;N. Kobayashi;J. Sawada;T. Terao;T. Nambara
本文介绍了一种测定11-脱氧皮质醇的单克隆抗体酶免疫分析法。用分泌抗11-脱氧皮质醇抗体的杂交瘤细胞在BALB/c小鼠腹水中产生抗11-脱氧皮质醇抗体。采用N-琥珀酰亚胺酯法对11-脱氧皮质醇进行酶标记。用β-半乳糖苷酶处理三种羧基化甾体的活化酯，得到酶标抗原。采用蛋白A-双抗体法分离免疫测定中的结合和游离组分。发现该单克隆抗体对由4-（2-羧基乙硫基）-11-脱氧皮质醇制备的同源酶标记抗原具有高亲和力，并且结合被分析物11-脱氧皮质醇抑制。在异源系统中使用的标记制备的11-脱氧皮质醇3-（O-羧甲基）肟和11-脱氧皮质醇21-半琥珀酸，较低和没有显着的免疫反应性，分别观察到。这些结果表明，单克隆抗体的抗原结合位点与11-脱氧皮质醇部分互补，并覆盖了该部分，该部分远离用于在免疫原制备中与载体连接的位置。同源分析系统显示出较高的灵敏度和特异性与使用氚作为示踪剂的放射免疫分析法相媲美。这些发现将有助于开发各种单克隆抗体免疫分析系统。
An enzyme immunoassay method using a monoclonal antibody for the determination of 11-deoxycortisol is described. The anti-11-deoxycortisol antibody was produced in ascites by inoculating antibody-secreting hybridoma cells into BALB/c mice. The enzyme labeling of 11-deoxycortisol was carried out by the N-succinimidyl ester method. The activated esters of three carboxylated steroids were treated with β-galactosidase to give enzyme-labeled antigens. A protein A-double antibody method was employed for separation of the bound and free fractions in the immunoassay. It.was found that the monoclonal antibody had a high affinity for a homologous enzyme-labeled antigen prepared from 4- (2-carboxyethylthio) -11-deoxycortisol and the binding was inhibited by the analyte, 11-deoxycortisol. In the heterologous systems using labels prepared from 11-deoxycortisol 3- (O-carboxymethyl) oxime and 11-deoxycortisol 21-hemisuccinate, a lower and no significant immunoreactivities were observed, respectively. These results indicate that the antigen-binding site of the monoclonal antibody is complementary to and covers the 11-deoxycortisol portion remote from the position used for attachment to, the carrier in the preparation of the immunogen. The homologous assay system showed high sensitivity and specificity comparable to those of the radioimmunoassay using tritium as a tracer. These findings should be helpful in the development of various immunoassay systems with the monoclonal antibody.