BLOOD SUGAR BY FOLIN- WU’S METHOD
- Principle OF BLOOD SUGAR BY FOLIN- WU’S METHOD
- normal range OF BLOOD SUGAR BY FOLIN- WU’S METHOD
- Specimen
- Reagents
- Requirements
- Procedure of BLOOD SUGAR BY FOLIN- WU’S METHOD
Principle of Blood sugar by Folin-Bu’s method
The protein free filtrate is heated with alkaline cupric tartarate by using special type of Folin-Wu tubes. Glucose (and other reducing substances) present in the filtrate reduce the cupric ions into cuprous ions. When phosphomolybdic acid reagent is added, it reacts with cuprous oxide to form molybdenum blue. Intensity of molybdenum blue is directly proportional to (the reducing substances and) glucose present in blood and it is measured photometrically and compared with a known standard.
Normal range
- a) Blood sugar (fasting): 80-120 mg/dl.
- b) Blood sugar: (P.P.): Up to 140 mg/dl. (2 Hrs. after lunch)
Specimen
2-3 ml of blood collected in a fluoride Vial.
Reagents
1. 2/3 N sulfuric acid.
2. 10 g/dl sodium tungstate.
3. Alkaline cupric tartarate
a) Sodium carbonate: 40 g
b) Tartaric acid: 7.5 g
Dissolve in about 800 ml distilled water ar add,
c) Copper sulphate: 4.5 g
Mix well and make final volume one liter. Store in a polyethylene bottle at room temperature (25 deg plus/minus 5 deg * C)
4 Phosphomolybdic acid
a) Molybdic acid: 35 g
b) Sodium tungstate: 5 g
c) 10 gm/di sodium hydroxide: 200 ml
d) Distilled water: 200 ml.
Mix and boil for one hour to remove ammonia.
Cool it to room temperature and add, Orthophosphoric acid: 125 ml.
e) Mix well and dilute to 500 ml by using distilled water. Store in a polyethylene bottle at room temperature (250c plus/minus 50 C)
5 .Glucose standard: 100 mg/dl (in saturated benzoic acid). Store in a polyethylene bottle at (2-8° C).
Requirements
1 Test tubes ( 15 x 125mm and 10 x100 mm)
2 Folin-Wu tubes
3 Pipettes: Graduated (2, 5 and 10 ml) and volumetric
(1.0 and 2.0 ml)
4 Funnels
5 Whatman no. 44 filter papers.
6 Water-bath
PROCEDURE of BLOOD SUGAR BY FOLIN- WU’S METHOD
A. Preparation of protein free filtrate Pipette in the tubes, labeled as follows-
| Distilled water, ml | 7.0 | 9.0 |
| Blood, ml | 1.0 | – |
| 100mg/dl glucose std. | – | 1.0 |
| 2/3 N sulphuric acid | 1.0 | – |
| 10 gm/dl sodium tungstate | 1.0 | – |
Mix thoroughly and keep at room temperature for 5 minutes. Filter the contents of test by using Whatman no. 44 filter paper.
B. Reduction of cupric ions
Pipette in the Folin-Wu tubes, labeled as follows:
| filtrate , ml | 2.0 | – | – |
| diluted glucose standard | – | 2.0 | – |
| distilled water | – | – | 2.0 |
| alkaline cupric tartarate | 2.0 | 2.0 | 2.0 |
Mix well and keep in a boiling water bath for exactly 8 minutes. Cool the tubes to room temperature.
C. Formation of molybdenum blue
Add 2.0 ml of phosphomolybdic acid in all the Folin-Wu tubes. Mix well.
Add distilled water up to 25 ml mark
Mix the contents thoroughly by inverting the tubes.
Keep for about 5 minutes.
Read the intensities of Test and standard against blank at 420 nm (violet filter).
calculation
O.D. of test /O.D. of std x 100
