Measurement of Blood urea

Method: -  Enzymatic colorimetric.

Principle: - Urea is hydrolyzed with the enzyme urease to produce ammonia & carbon dioxide the ammonium ions react with hypochlorite and salicylate to give green dye. The intensity of the color is measured by colorimeter.

 Urea+H2O   --------------> 2NH3+CO2

NH3+Hypochlorite+Salicylate   ® Green dye.

Apparatus required:

     

       ®   Colorimeter

       ®   Test tubes with rack

       ®   Pipettes

       ®   Timer + Marker

       ®   Blood collection set

Reagents required:-

  Ø Reagent 1

  Ø Reagent 2

  Ø Urease Enzyme

  Ø Standard (80 mg/dl)

Sample/Specimen:

     ®  Serum or Haparinized plasma which must be free from haemolysis.

Procedure/Assay:-

    Ø Wavelength- 578 nm(570-600 nm)

    Ø Cuvette-1cm light path

    Ø Temperature-25°C

Pipette into test tubes as follows:

Mix incubation for 5 min. at 20-25°C or for 3 min. at 37°C

Mix incubation for 10 min. at 20-25°C or for 5 min. at 37°C.measure the absorbance of the sample and the standard against the reagent blank within 60 min.

Calculation:-          

Test Result:- Urea reference range:10-50 mg/dl or 1.7-8.3 mmol/l