Aubin P, Le Brun G, Moldovan F, Villette J M, Créminon C, Dumas J, Homyrda L, Soliman H, Azizi M, Fiet J
Laboratoire de Biologie Hormonale, Hôpital Saint-Louis, Paris, France.
Clin Chem. 1997 Jan;43(1):64-70.
A sandwich-type enzyme immunoassay has been developed for measuring human big endothelin-1 (big ET-1) in human plasma and supernatant fluids from human cell cultures. Big ET-1 is the precursor of endothelin 1 (ET-1), the most potent vasoconstrictor known. A rabbit antibody raised against the big ET-1 COOH-terminus fragment was used as an immobilized antibody (anti-P16). The Fab' fragment of a monoclonal antibody (1B3) raised against the ET-1 loop fragment was used as the enzyme-labeled antibody, after being coupled to acetylcholinesterase. The lowest detectable value in the assay was 1.2 pg/mL (0.12 pg/well). The assay was highly specific for big ET-1, demonstrating no cross-reactivity with ET-1, <0.4% cross-reactivity with big endothelin-2 (big ET-2), and <0.1% with big endothelin-3 (big ET-3). We used this assay to evaluate the effect of two different postural positions (supine and standing) on plasma big ET-1 concentrations in 11 male and 11 female healthy subjects. Data analysis revealed that neither sex nor body position influenced plasma big ET-1 concentrations. This assay should thus permit the detection of possible variations in plasma concentrations of big ET-1 in certain pathologies and, in association with ET-1 assay, make possible in vitro study of endothelin-converting enzyme activity in cell models. Such studies could clarify the physiological and clinical roles of this family of peptides.
已开发出一种夹心型酶免疫测定法，用于检测人血浆和人细胞培养上清液中的人 big 内皮素 -1（big ET-1）。Big ET-1 是内皮素 1（ET-1）的前体，ET-1 是已知最强效的血管收缩剂。一种针对 big ET-1 羧基末端片段产生的兔抗体用作固定化抗体（抗 -P16）。一种针对 ET-1 环片段产生的单克隆抗体（1B3）的 Fab' 片段在与乙酰胆碱酯酶偶联后用作酶标记抗体。该测定法的最低检测值为 1.2 pg/mL（0.12 pg/孔）。该测定法对 big ET-1 具有高度特异性，与 ET-1 无交叉反应，与 big 内皮素 -2（big ET-2）的交叉反应性<0.4%，与 big 内皮素 -3（big ET-3）的交叉反应性<0.1%。我们使用该测定法评估了两种不同体位（仰卧和站立）对 11 名男性和 11 名女性健康受试者血浆 big ET-1 浓度的影响。数据分析显示，性别和体位均不影响血浆 big ET-1 浓度。因此，该测定法应能检测某些病理状态下血浆 big ET-1 浓度的可能变化，并与 ET-1 测定法相结合，在细胞模型中对内皮素转化酶活性进行体外研究。此类研究可阐明该肽家族的生理和临床作用。
