Diagnostic Laboratory Section

ESTIMATION OF PROTEIN AND ALBUMIN  

Protein

Principle (Direct biuret method)

• Colorimetric determination of total protein based on the principle of the Biuret reaction (copper salt in an alkaline medium). Protein in plasma or serum sample forms a blue colored complex when treated with cupric ions in alkaline solution. The intensity of the blue colour is proportional to the protein concentration.

Reagent composition

• Potassium iodide - 6 mmol/L
• Potassium sodium tartarate  - 21 mmol/L
• Copper Sulphate -  6 mmol/L
• Sodium hydroxide  - 58 mmol/L

Laboratory procedure

	Blank	Standard	Sample
Reagent	1000μL	1000μL	1000μL
Standard	--	20 μL	--
Sample	--	--	20μL
Mix and incubate for 10 min. at 37 ° C. Measure the absorbance of standard and sample against reagent blank.

Calculation 

 Total Protein Conc. (g/dl) = (Absorbance of Sample / Absorbance of standard ) x 6

           Total Protein Concentration in standard = 6 g/dl.

Albumin

Method of Estimation : Bromocresol green method.

Principle

• The reaction between albumin from serum or plasma and the dye bromocresol- green produces a change in colour that is proportional to the albumin concentration.

Reagent composition

• Succinate buffer, (pH 4.20) 75 mmol/ L
• Bromocresol green 0.14 g/L

Laboratory procedure

	Blank	Standard	Sample
Reagent	1000μL	1000μL	1000μL
Standard	--	10 μL	--
Sample	--	--	10μL
Mix and incubate for 1 minute. Measure the absorbance of standard and sample against reagent blank.

Calculation

• Albumin Conc. (g /dl) = (Absorbance of sample / Absorbance of standard ) x 3
• Albumin standard concentration =  g/ dL