Determination of diadenosine 5',5''',-P1,P4-tetraphosphate levels in cultured mammalian cells.
Determination of diadenosine 5',5''',-P1,P4-tetraphosphate levels in cultured mammalian cells.
复制标题
测定培养的哺乳动物细胞中的二腺苷 5,5,-P1,P4-四磷酸水平。
DOI：
10.1016/0003-2697(84)90070-8
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发表时间：
1984
影响因子：
2.9
通讯作者：
Jacobson,MK
中科院分区：
文献类型：
--
作者：
Baker,JC;Jacobson,MK
本文报道了一种测定哺乳动物细胞中二腺苷5′，5 ′-P1，P4-四磷酸（Ap 4A）含量的简便方法。通过使用0.1 N NaOH溶解细胞单层来快速提取Ap 4A。通过选择性吸附到高度特异性的硼酸酯亲和树脂上，在一个步骤中将其从大量细胞组分中分离出来。通过在碱性磷酸酶和毒液磷酸二酯酶存在下进行的荧光素酶生物发光测定来定量亚皮摩尔量。通过高效液相色谱法确定了该测定法对培养小鼠细胞中Ap 4A的选择性。该方法允许常规测量来自单个细胞培养皿的亚皮摩尔量的Ap 4A。
A simple method for measuring the cellular content of diadenosine 5′,5‴-P1,P4-tetraphosphate (Ap4A) in cultured mammalian cells is described. Ap4A was rapidly extracted by dissolving cell monolayers using 0.1 n NaOH. It was separated from the bulk of cellular components in a single step by selective adsorption to a highly specific boronate affinity resin. Subpicomole amounts were quantified by a luciferin-luciferase bioluminescence assay performed in the presence of alkaline phosphatase and venom phosphodiesterase. The selectivity of this assay for Ap4A in cultured mouse cells was established by high-performance liquid chromatography. This method allows the routine measurement of subpicomole amounts of Ap4A derived from a single dish of cells.