3-磷酸甘油醛脱氢酶活性检测方法的改良及应用

摘要

目的 改良3-磷酸甘油醛脱氢酶(GAPDH)活性检测方法,并探讨该方法的应用.方法在经典血清GAPDH检测方法基础上,根据该酶的组织分布特点,将血清样本改为全血样本,并对样本及检测条件作了进一步的改良.在方法学研究的基础上,运用改良后的方法分别检测正常鹌鹑、高嘌呤饮食诱导的模型鹌鹑、正常大鼠及高果糖饮食诱导的模型大鼠全血GAPDH的活性.结果 全血与血清GAPDH酶促反应时间曲线类似,变异系数均小于10%.方法 学应用结果显示,与正常鹌鹑比,高嘌呤饮食诱导的模型鹌鹑在高尿酸血症及其合并脂、糖代谢紊乱状态下(60～140 d)全血GAPDH活性显著降低;与正常大鼠比,高果糖饮食诱导的模型大鼠在高三酰甘油血症及其并发尿酸、糖代谢紊乱下(7～28 d),全血GAPDH活性显著降低.结论 改良方法血样容易采集,样品用量少,操作简便、重复性较好,经济实用,易于推广应用.%Objective To improve the determination method of glyceraldehyde-3-phosphate dehydrogenase(GAPDH) activity and explore its application. Methods The traditional determination method of GADPH activity was amended by changing serum sample as whole blood sample and reforming the related determination condition, according to GAPDH distribution in the tissue. And then GAPDH activity was detected by newly constructed method on quail and rat models. Results The time curve of whole blood GAPDH activity was similar to serum curve,and coefficient of variation of the tow methods were both less than 10%. The activity of GAPDH was decreased in quails model group,induced by high purine diet, with hyperuricemia and combined with metabolic disturbance of blood fat and glucose(60 - 140 d) ,and was also decreased in rats model group,induced by high fructose diet, with hy-pertriacylglycerolemia and combined with metabolic disturbance of blood uric acid and glucose(7 - 28 d). Conclusion This newly constructed method could have several advantages as easy collection of sample, less usage amount of sample, good reproducibility and economics.