Rasmussen P H, Plum I G, Bruun N E, Dige-Petersen H, Hedner T, Hedner J, Giese J
Department of Clinical Physiology and Nuclear Medicine, Glostrup Hospital, University of Copenhagen, Denmark.
Blood Press. 1992 Oct;1(3):181-6. doi: 10.3109/08037059209077515.
A specific and sensitive radioimmunoassay (RIA) for determination of endothelin-1 (ET-1) in human plasma has been developed. Antibodies were raised in rabbits using synthetic ET-1 conjugated to thyroglobulin as immunogen. The antibodies obtained were used at a final dilution of 1:300,000 yielding maximum binding of 61.7 +/- 3.0% (mean +/- 1 SD, n = 20) of 125I-ET-1. The ID50 (inhibitory dose 50%) was 4.5 +/- 0.6 fmol/100 microliters (mean +/- 1 SD, n = 20). The sensitivity of the RIA was 0.33 fmol/100 microliters standard solution. No cross reactivity was observed with endothelin-3, big-endothelin-1, atrial natriuretic factor, angiotensin I or angiotensin II. The cross-reactivity with endothelin-2 was 100%. Endothelin was extracted from acidified plasma with Sep-pak C18 cartridges and recovery of ET-1 added to normal plasma was 70.9 +/- 10.3% (mean +/- 1 SD, n = 12). The concentration of ET-1 in plasma from normal subjects was 1.5 +/- 0.4 pmol/l (mean +/- 1 SD, n = 11) ranging from 1.0 to 2.2 pmol/1. Extracts of normal human plasma subjected to high performance liquid chromatography on a reverse phase C18 column showed one peak of immunoreactivity co-eluting with the standard for ET-1. From these data it is concluded that the immunoreactive material measured in normal plasma with the present RIA is identical to ET-1.
已开发出一种用于测定人血浆中内皮素 -1（ET -1）的特异性和灵敏性放射免疫分析（RIA）方法。以与甲状腺球蛋白偶联的合成ET -1作为免疫原在兔体内产生抗体。所获得的抗体以1:300,000的终浓度稀释使用，对125I -ET -1的最大结合率为61.7±3.0%（平均值±1标准差，n = 20）。半数抑制剂量（ID50）为4.5±0.6 fmol/100微升（平均值±1标准差，n = 20）。该RIA的灵敏度为0.33 fmol/100微升标准溶液。未观察到与内皮素 -3、大内皮素 -1、心房利钠因子、血管紧张素I或血管紧张素II的交叉反应。与内皮素 -2的交叉反应率为100%。用Sep - pak C18柱从酸化血浆中提取内皮素，添加到正常血浆中的ET -1回收率为70.9±10.3%（平均值±1标准差，n = 12）。正常受试者血浆中ET -1的浓度为1.5±0.4 pmol/L（平均值±1标准差，n = 11），范围为1.0至2.2 pmol/L。正常人血浆提取物在反相C18柱上进行高效液相色谱分析，显示一个免疫反应峰与ET -1标准品共洗脱。从这些数据可以得出结论，用本RIA法在正常血浆中测得的免疫反应性物质与ET -1相同。
