Phosphate Assay
Categorized: Biochemical Assays
Stock Solution:
0.001 M Phosphate Solution
use 100, 200, 300, 400, 500 µl and make up each sample to 500 µl final volume.
Reagents:
- 10% Magnesium Nitrate in ethanol.
- 1 M HCl.
- 10% Ascorbic Acid (1 gm/10 ml).
- Ammonium Molybdate, .42% in 1 N Sulphuric Acid
(.42 gm + 2.66 ml c/H2SO4 made up into 100 ml)
Method:
- Weigh 0.5 mg LPS into Pyrex tube.
- Add 0.05 ml 10% Magnesium Nitrate in ethanol.
- Evaporate to dryness on flame.
- Cool. Add 0.3 ml 1 N HCl. Heat at 100oC for 15 min. in a boiling water bath.
- Add 0.7 ml Ascorbic/Molybdate (1 part to 6 parts freshly mixed).
- Keep at 45oC for 20 min.
- Read absorbance at 820 nm against reagent blank. (absorbence of 0.240 for 0.01 µmol phosphate).
* Ascorbic Acid is stable at 2 – 4oC for approx. 7 weeks.
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