分光光度法测定红细胞嘌呤核苷磷酸化酶

Determination of purine nueleoside phosphorylase in erythocytes with spectrophotometric assay

应用Trinder反应建立终点分光光度法测定红细胞嘌呤核苷磷酸化酶(PNP)活性,操作简单、快速、灵敏、准确。对酶反应最佳条件进行了实验探讨,底物Km为64mm,最佳PH7.5,延滞期1分钟30秒,1.5～8分钟酶活性与反应时间成比例。用3.5mol/L硫酸铵溶液终止反应,效果好,显色稳定。批内CV2.51％～3.15％,批间CV2.69％～3.28％。35例正常人红细胞PNP活性为13.66±1.73u/ml红细胞。

It was described that spectrophometric assay was established for determination of purine nueleoside phosphorylase (PNP) in erytrocytes by Trinder. It is simple, rapid, sensitive and accurate method. Optium reaction conditions for the activity of PNP were studied. The results showed that Km value was 84 um, and that PH was 7.50 in the substrated. The delay period was 1.5 min. The relationship between the activity of PNP and reaction time was linear up from 1.5 to 8 min. The PNP activity was inhibitored effectively by 3.5 mol/L(NH_4)_2SO_4 and color is stability. Withinrun precision CV was 2.51～3.15% and betweenrun precision (CV) were 2.69～3.28%. The mean eryocyte PNP activity was 13.66±1.73U/ml in 35 healthy subjects.