Blood proteins are precipitated by zinc hydroxide. The filtrate is heated with
alkalinecopper reagent and the reduced Cu formed is treated with arsenomolybdate
reagent, resulting in the formation of violet, which is read in the
- 5% ZnSO4 solution
- 0.34-N barium hydroxide
These 2 solutions should be adjusted so that 5 mL ZnSO4 require
Ba(OH)2 for complete neutralization.
- Alkaline copper reagent:
Solution A: 25-gm anhydrous Na2S2O3, 25-gm Rochelle salt, 20-gm NaHCO3,
and 200-gm anhydrous Na2SO4 were dissolved in about 800 mL of water
and diluted to 1 liter. The solution was stored at room temperature and
never dipped below 20°C. It was filtered before use if any sediment was
Solution B: 15% CuSO4, 5H2O containing 1 or 2 drops of concentrated
H2SO4. On the day of use, 25 parts of solution A and part of solution B
were mixed. That was the alkaline copper reagent.
- Arsenomolybdate color reagent: Ammonium molybdate 25 gms was
dissolved in 450 mL H2O. 21 mL concentrated H2SO4 was added and
mixed. Disodium orthoarsenate (Na2H, ASO4, 7H2O) 3 gms was dissolved
in 25 mL H2O and added with stirring to the acidified molybdate solution.
It was then placed in an incubator at 37°C for 24–48 hours and stored in
a glass-stoppered brown bottle.
- Standard glucose solution: (stock glucose solution)
Exactly 0.1 gm of anhydrous pure glucose was dissolved in 10–15 mL
0.2% benzoic acid and diluted to 100 mL with benzoid acid solution.
Three working standards were prepared by diluting 0.5, 1.0, and 2.00 mL
of the stock solution to 100 mL with benzoic acid. These solutions in benzoic
acid kept indefinitely at room temperature.
Working standard: 5 mL made up to 100 mL yields 50 mg/M.
Into a test tube containing 3.5 mL of water, 0.1 mL of blood was introduced
through a clean dry micropipette and mixed well. To this tube, 0.2 mL of 0.3
N Ba(OH)2 was added after the mixture turned brown. 0.2 mL ZnSO4 was
added and mixed. After 10–15 minutes, the mixture was filtered through a
Whatman No. 1 filter paper.
Into 2 separate test tubes, 1 mL aliquot of the filtrate was transferred and
1 mL alkaline Cu reagent was then added. These tubes were covered with glass
and placed in a boiling water bath for 20 minutes. The tubes were then cooled
under running water, 1 mL arsenomolybdate reagent was added and the solution
dilute to 25 mL with H2O simultaneously standards in 0.2 to 1.0 mL range were
prepared and a reagent blank were similarly prepared.
The intensity of color produced was red at 680 mm. The color was stable
and reading may be taken at convenience.
The concentration of blood glucose in given sample is .............. mg/mL