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3PLE

ref: http://doi.org/10.1194/jlr.D090795

Protocol​

Performed at room temperature (including centrifugation) to maintain consistent solubility and phase separation.

For ~30mg of tissue (liver, brain and white adipose tissue) or xxxxxx

  1. Add 300μl of hexanes, 300μl of methyl acetate, 225μl of acetonitrile, and 300μl of water to the glass tube containing the sample.
  2. Vortex for 10 min.
  3. Centrifuge at 3000xg for 5 min.
  4. Collect the upper phase and the middle phase into separate glass tubes.
  5. Re-extraction of the middle phase.
  6. Add 300μl of hexane to the middle phase.
  7. Vortex for 10 min.
  8. Centrifuge at 3000xg for 5 min.
  9. Collet the bottom phase to a fresh glass tube.
  10. Dry under nitrogen.

The lipid fractionation is based on polarity. The upper phase is enriched in neutral lipids while the middle phase contains the glycerophospholipids and other polar lipid species.