Measurement of Phosphoinositide 3-Kinase Products in Cultured Mammalian Cells by HPLC

  • Frank T. CookeEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 645)


The phosphoinositide 3-kinase (PI3K) family catalyses the addition of a phosphate group to the D-3 position of polyphosphoinositides (PPIn). Since the discovery in the late 80s that phosphatidylinositol is phosphorylated in the D-3 position in eukaryotic cells, there has been an explosion of interest in these PPIn. Although the four D-3 PPIn (phosphatidylinositol 3-phophate (PtdIns3P), PtdIns(3,4)P 2, PtdIns(3,5)P 2, and PtdIns(3,4,5)P 3) represent only a small proportion of PPIn, production of D-3 PPIn is required for an ever-increasing number of processes. Measurement of the PPIn levels in intact cells cultured cells has been vital to our understanding of the metabolism and function of these important signalling molecules; methods are described herein that allow measurement of PPIn levels in cultured cells, with emphasis on the 3-OH PPIn.

Key words

PI3K Phosphoinositides HPLC Isotopic labelling 



bovine serum albumin


counts per minute


disintegrations per minute




fatty acid-free




HEPES buffered balanced salts solution


high performance liquid chromatography






phosphoinositide 3-kinase




inorganic PO4


tetrabutyl ammonium hydrogen sulphate


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Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  1. 1.Department of Structural and Molecular BiologyUniversity College LondonLondonUK

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