| Contributor: |
The Laboratory of Giovanna Ferro-Luzzi Ames at the University of California, Berkeley
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| This protocol measures the amount of carbohydrate in a sample. Even a 10-fold excess of protein will not interfere with the quantification of carbohydrate in this assay. |
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1. Start with the sample dissolved in ddH2O (total volume up to 1 ml) in a fraction collector test tube.
2. Add 2 ml of 4% Phenol.
3. Quickly add 5 ml of Concentrated Sulfuric Acid onto the liquid surface (CAUTION! see Hint #1).
4. Mix tube contents immediately by vortexing. The solution gets very hot.
5. Read the absorbance at 490 nm in a spectrophotometer approximately 30 min later (see Hint #2).
6. Read the absorbance at 490 nm of a set of Glucose Standards ranging from 0 μg to 100 μg. Construct a standard curve.
7. Compare the absorbance values of the samples against a standard curve generated with Glucose to calculate the Glucose concentration.
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| 4% Phenol |
| 4% (v/v) Phenol in ddH2O (CAUTION! see Hint #1)
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Glucose Standards
Sulfuric Acid, Concentrated
Phenol
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1. CAUTION! This substance is a biohazard. Consult this agent's MSDS for proper handling instructions.
2. The color is stable for a least this amount of time and will reach a development plateau within 30 min.
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