Lipid Phosphate Determination

 David K. Perry for info

- Evaporate solvent from lipids in CHCl₃ phase under N₂ or in a speed vac. concentrator (use Pyrex test tubes)

- Prepare a standard curve of 0-40 nmoles of Pi using Na- or K-phosphate

- Add to each tube 0.6 ml of 10N H₂SO₄: 70% HClO₄: H₂O (9:1:40)

- Incubate overnight at 160° C

- Add 0.9 ml H₂O and vortex

- Add 0.5 ml of 0.9% ammonium molybdate and vortex

- Add 0.2 ml of 9.0% ascorbic acid (made fresh) and vortex

- Incubate for 30 min at 45° C

- Read absorbance at 820 nm

- Calculate nmoles of phosphate based on slope of standard curve

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