血清糖基化磷脂酰肌醇特异性磷脂酶D的检测

Methodological study on the assay of glycosylphosphatidylinositol specific phospholipase D activity in serum

利用自制的、具完整糖基化磷脂酰肌醇（ＧＰＩ）结构的胎盘型碱性磷酸酶（ＰＬＡＰ）做底物，通过聚乙二醇／葡聚糖分相系统定性，ＴＸ１１４分相定量，初步建立了检测人血清中ＧＰＩ特异性磷脂酶Ｄ（ＧＰＩＰＬＤ）的方法，并对酶促反应的某些动力学性质进行了探讨。该方法重现性好（ＣＶ＝３．６％），灵敏度高（血清用量２μｌ即可），且经济、实用，有一定的推广价值。用该方法检测１００名正常人血清，表明其ＧＰＩＰＬＤ的活性范围为３０％～５０％（用转化底物的百分率表示）。

A method for the assay of glycosylphosphatidylinositol specific phospholipase D(GPI PLD) activity in human serum was established by using glycosylphosphatidylinositol(GPI) anchored placental alkaline phosphatase(PLAP) as a substrate. Serum GPI PLD activity was determined both qualitatively and quantitatively by PEG/dextran and triton X 114 partitioning respectively, and some kinetic properties of its enzymatic reaction were also studied. This method revealed a satisfactory reproductivity(CV=3.6%) and a high sensitivty(2μl serum was sufficient). In addition, it is practical, economical, and feasible to be widely applied. The activities of GPI PLD measured by this method in 100 normal subjects ranged from 30% to 50%(in terms of the percentage of converting GPI anchored PLAP substrate to the anchor free product).