PMID: 2118761Aug 15, 1990Paper

Isolation and enzymic assay of choline and phosphocholine present in cell extracts with picomole sensitivity

The Biochemical Journal
J J MurrayD A Kennerly

Abstract

Increasing interest in receptor-regulated phospholipase C and phospholipase D hydrolysis of cellular phosphatidylcholine motivates the development of a sensitive and simple assay for the water-soluble hydrolytic products of these reactions, phosphocholine and choline respectively. Choline was partially purified from the methanol/water upper phase of a Bligh & Dyer extract by ion-pair extraction using sodium tetraphenylboron, and the mass of choline was determined by a radioenzymic assay using choline kinase and [32P]ATP. After removal of choline from the upper phase, the mass of residual phosphocholine was determined by converting it into choline by using alkaline phosphatase, followed by radioactive phosphorylation. In addition to excellent sensitivity (5 pmol for choline and 10 pmol for phosphocholine), these assays demonstrated little mutual interference (phosphocholine----choline = 0%; choline----phosphocholine = 5%), were extremely reproducible (average S.E.M. of 3.5% for choline and 2.9% for phosphocholine), and were simple to perform with instrumentation typically available in most laboratories. In addition, the ability to apply the extraction technique to the upper phase of Bligh & Dyer extracts permitted simple analysi...Continue Reading

Citations

Jul 31, 2012·Analytical Biochemistry·Le Duy DoSaida Mebarek
Oct 26, 2011·Journal of Lipid Research·Aditya BansalTimothy R DeGrado
Sep 6, 2008·Analytical and Bioanalytical Chemistry·Tatyana V BarkhimerL M Viranga Tillekeratne
Nov 5, 1997·Analytical Biochemistry·A J MorrisJ Engebrecht
Dec 17, 2002·Journal of Pharmacological and Toxicological Methods·S MuraiT Kawaguchi
Mar 1, 1995·Metabolism: Clinical and Experimental·K P Boggs, M G Buse

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