PHOTOMETRIC MICROTITER ASSAY OF INORGANIC-PHOSPHATE IN THE PRESENCE OF ACID-LABILE ORGANIC-PHOSPHATES
PHOTOMETRIC MICROTITER ASSAY OF INORGANIC-PHOSPHATE IN THE PRESENCE OF ACID-LABILE ORGANIC-PHOSPHATES
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DOI：
10.1006/abio.1995.1453
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发表时间：
1995-09-01
影响因子：
2.9
通讯作者：
PALM, D
中科院分区：
文献类型：
--
作者：
DRUECKES, P;SCHINZEL, R;PALM, D
一个简单而快速的比色微量测定无机磷酸盐在温和的酸性pH值范围内，使我们能够进行广泛的酶动力学研究，即使在高浓度的酸不稳定底物的存在下。在96孔微量滴定板中使用30 μ l含有0.5 - 100 nmol P-i的样品进行测定。与现有的微量测定相比，颜色形成在更宽的磷酸盐浓度和反应条件范围内呈线性。使用计算机辅助微量滴定板读数器，数据可以直接传输到动力学分析软件。上述测定用于在包括在单个96孔微量滴定板上以可变底物浓度和pH筛选底物和抑制剂的条件下，同时测定糖原合成方向上大肠杆菌麦芽糖糊精磷酸化酶催化反应的多达12个速率。(C)出版社：Academic Press
A simple and rapid colorimetric microassay for inorganic phosphate in the mild acid pH range has enabled us to perform extensive enzyme kinetic studies even in the presence of high concentrations of acid-labile substrates. The assay is performed in a 96-well microtiter plate using 30-mu l samples containing between 0.5 and 100 nmol P-i. Compared to existing microassays, color formation is linear over a much wider range of phosphate concentrations and reaction conditions. Using a computer-assisted microtiter plate reader, the data can be directly transferred to kinetic analysis software. The above assay was used to determine simultaneously up to 12 rates of the Escherichia coli maltodextrin phosphorylase-catalyzed reaction in the direction of glycogen synthesis under conditions which include screening of substrates and inhibitors at variable substrate concentrations and pH on a single 96-well microtiter plate. (C) 1995 Academic Press, Inc.