Measurement of phospholipid metabolism in intact neutrophils

Methods Mol Biol. 2014:1124:89-105. doi: 10.1007/978-1-62703-845-4_7.

Abstract

Phospholipid-metabolizing enzymes are important participants in neutrophil signal transduction pathways. The methods discussed herein describe assays for assessing the activities of phospholipase A2 (PLA2), phospholipase C (PLC), phospholipase D (PLD), and phosphoinositide 3-OH-kinase in intact neutrophils. PLA2 activity is measured as the release of radiolabeled arachidonic acid. PLC activity is measured as the accumulation of inositol 1,4,5-trisphosphate (IP3), a water-soluble product, using a commercially available radioreceptor assay kit. PLD activity is measured as the appearance of its radiolabeled products, phosphatidic acid and phosphatidylethanol. PI3-K activity is measured as the appearance of its radiolabeled product, phosphatidylinositol-3,4,5-trisphosphate.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Chromatography, Thin Layer / methods
  • Enzyme Activation
  • Group IV Phospholipases A2 / metabolism
  • Humans
  • Lipid Metabolism*
  • Neutrophils / metabolism*
  • Phosphatidylinositol 3-Kinases / metabolism
  • Phospholipase D / metabolism
  • Phospholipids / metabolism*
  • Signal Transduction
  • Type C Phospholipases / metabolism

Substances

  • Phospholipids
  • Phosphatidylinositol 3-Kinases
  • Group IV Phospholipases A2
  • Type C Phospholipases
  • Phospholipase D