Ohta H, Tsuji T, Asai S, Tanizaki S, Sasakura K, Teraoka H, Kitamura K, Kangawa K
R&D and Manufacturing Department for Diagnostics, Shionogi & Co., Ltd., Settsu, Osaka, Japan.
Clin Chim Acta. 1999 Sep;287(1-2):131-43. doi: 10.1016/s0009-8981(99)00128-x.
We developed a one-step two-site immunoradiometric assay (IRMA) using two kinds of monoclonal antibodies, which enables us to directly measure the entire molecules of adrenomedullin (AM) (the sum of mature-type AM (abbreviated, m-AM) amidated at the C-terminus and Gly-extended non-amidated AM) in human plasma using a small amount of sample (100 microl) without prior extraction. The detection limit of this assay was 0.5 pmol/l for a 100-microl sample. Intra- and inter-assay precisions were 3.4-7.3% and 5.8-7.6%, respectively. The dilution curves of plasma samples showed good linearity and analytical recovery was 89-118%. The mean total AM in plasma of healthy subjects was 9.00+/-2.13 pmol/l, whereas m-AM was 1.05+/-0.24 pmol/l. This method, together with our previously reported simplified method to specifically measure m-AM (Ohta et al., Clin Chem 1999;45:244-251), allows facile estimation of the plasma concentration of AM-Gly by subtracting m-AM from the total AM measured by the procedure described in this paper. We were able to show that the concentration of total AM in patients with sepsis was markedly higher than that in the healthy controls and that the ratios of m-AM/total AM were significantly different between the controls and patients.
我们使用两种单克隆抗体开发了一种一步双位点免疫放射分析（IRMA）方法，该方法使我们能够直接测量人血浆中肾上腺髓质素（AM）的完整分子（C末端酰胺化的成熟型AM（缩写为m-AM）和Gly延伸的非酰胺化AM的总和），只需少量样本（100微升），无需预先提取。该分析方法对100微升样本的检测限为0.5皮摩尔/升。批内和批间精密度分别为3.4-7.3%和5.8-7.6%。血浆样本的稀释曲线显示出良好的线性，分析回收率为89-118%。健康受试者血浆中总AM的平均值为9.00±2.13皮摩尔/升，而m-AM为1.05±0.24皮摩尔/升。该方法与我们之前报道的特异性测量m-AM的简化方法（Ohta等人，《临床化学》1999年；45:244-251）一起，通过从本文所述方法测量的总AM中减去m-AM，可以轻松估算血浆中AM-Gly的浓度。我们能够证明，脓毒症患者的总AM浓度明显高于健康对照组，并且对照组和患者之间m-AM/总AM的比率存在显著差异。
