Matsushita M, Irino T, Komoda T, Sakagishi Y
Department of Clinical Chemistry, Saitama College of Health, Urawa, Japan.
Clin Chim Acta. 1993 Jul 16;216(1-2):103-11. doi: 10.1016/0009-8981(93)90143-r.
A method of protein determination has been developed which combines the biuret reaction and the copper(I)-bathocuproine chelate reaction. Protein in the specimen forms a Cu(2+)-protein chelate complex (biuret reaction) during the first step. Excess Cu2+ is reduced to Cu+ by ascrobic acid, allowing the Cu+ to form a Cu(+)-bathocuproine chelate complex during the second step. The amount of Cu(+)-bathocuproine chelate complex formed is inversely proportional to the protein concentration. The sensitivity (epsilon = 1.4 x 10(6) 1.mol-1.cm-1 against human albumin) of this method was higher than that of the original Lowry (9.8 x 10(5)), pyrogallol red (1.0 x 10(6)) and commercially available Coomassie Brilliant Blue G.250 methods (6.7 x 10(5)). The color intensities of human gamma-globulin, human globulin (fractions IV-1 and IV-4), bovine albumin, egg albumin and horse gamma-globulin against human albumin (100%) ranged from 92 to 101%. The results obtained with the present method (y) correlated well with those determined by the biuret method (r = 0.998, y = 0.98 chi - 0.002, x = 1.31, y = 1.29 g/l) in 30 diluted sera. These results confirm that this assay is similar in sensitivity to the original Lowry method, is rapid and has similar reactivity to each of the various proteins in biological fluids.
已开发出一种蛋白质测定方法，该方法结合了双缩脲反应和铜（I）- bathocuproine螯合反应。在第一步中，标本中的蛋白质形成Cu（2 +）-蛋白质螯合复合物（双缩脲反应）。过量的Cu2 +被抗坏血酸还原为Cu +，从而在第二步中使Cu +形成Cu（+）- bathocuproine螯合复合物。形成的Cu（+）- bathocuproine螯合复合物的量与蛋白质浓度成反比。该方法的灵敏度（针对人白蛋白，ε= 1.4 x 10（6）1.mol-1.cm-1）高于原始的Lowry法（9.8 x 1憨绩封啃莩救凤寻脯默0（5））、邻苯三酚红法（1.0 x 10（6））和市售的考马斯亮蓝G.250法（6.7 x 10（5））。人γ-球蛋白、人球蛋白（IV - 1和IV - 4组分）、牛白蛋白、卵白蛋白和马γ-球蛋白相对于人白蛋白（100%）的颜色强度范围为92%至101%。在30份稀释血清中，用本方法获得的结果（y）与双缩脲法测定的结果相关性良好（r = 0.998，y = 0.98χ - 0.002，x = 1.31，y = 1.29 g/l）。这些结果证实，该测定法在灵敏度上与原始的Lowry法相似，速度快，并且对生物流体中的各种蛋白质具有相似的反应性。
