Asai K, Asai M
Clin Chim Acta. 1984 Dec 29;144(2-3):163-71. doi: 10.1016/0009-8981(84)90050-0.
A colorimetric assay for plasma antithrombin III (ATIII) has been developed, using a new synthetic peptide substrate, PS-915 (H-D-Phe-Pro-Arg-CHA). PS-915 is freely water-soluble and its aqueous solution is stable. It is highly specific for thrombin and its Km value is about the same as that of S-2238. By enzyme hydrolysis, the new substrate liberates 3-carboxy-4-hydroxyaniline (CHA), which turns blue in color due to the complex formation with added alkaline-pentacyanoammine ferroate. The assay is a three-stage kinetic, one-point procedure, based on the method of antithrombin (heparin cofactor activity) determination. Since absorption measurements of the CHA are usually obtained at 700 nm, no blank correction is necessary even when hyperbilirubinemic, lipemic and/or hemolyzed plasma are used. The calibration curve for ATIII is linear over the range of 0-175% of normal and has a high degree of reproducibility. The results obtained by this method are well correlated with data obtained by the commonly used chromogenic assay or by radial immunodiffusion.
已开发出一种使用新型合成肽底物PS-915（H-D-苯丙氨酸-脯氨酸-精氨酸-环己基氨基甲酸酯）检测血浆抗凝血酶III（ATIII）的比色测定法。PS-915可自由溶于水，其水溶液稳定。它对凝血酶具有高度特异性，其Km值与S-2238的Km值大致相同。通过酶水解，新底物释放出3-羧基-4-羟基苯胺（CHA），由于与添加的碱性五氰合氨铁酸盐形成络合物，CHA会变成蓝色。该测定法是基于抗凝血酶（肝素辅因子活性）测定方法的三步动力学单点程序。由于CHA的吸光度测量通常在700nm处进行，因此即使使用高胆红素血症、脂血和/或溶血血浆也无需进行空白校正。ATIII的校准曲线在正常范围的0-175%内呈线性，具有高度的重现性。通过该方法获得的结果与常用的显色测定法或放射免疫扩散法获得的数据高度相关。
