Serum biochemical parametersTOTAL Protein assay

Biochemistry Clinical practiceCLS 432

Dr. Samah KotbLecturer of Biochemistry

2015

What do you know about liver function

• 1. Total and conjugated bilirubin • 2. ALT• 3. AST• 4. Total protein• 5. GGT• 6. ALP • 7. 5 -nucleotidase′• 8. Albumin• 9. prothrombin time

Liver Function tests

• Total protein was estimated according to the method of (Burtis A and Koller A ., 1999) , Colorimetric method.

Principle

• Proteins give an intensive violet –blue complex with copper salts in an alkaline medium. Iodide is included as an antioxidant.

Principle

• The intensity of the color formed is proportional to the total protein concentration in the sample.

• Total protein Standard : Bovine serum albumin standard (7g/dL)(70 g/L).

• Biuret reagent: R1

• Sodium Potassium tartrate (21 mmol/L)

• Sodium hydroxide (0.75 mol/L)

• Potassium iodide (6 mmol/L)

• Cupric Sulfate (6 mmol/L)

Reagents

• 1- 1 ml of Biuret reagent was added into both unknown test and control tubes, then 0.02 ml of serum were added to unknown.

Procedure

• 2- 0.02 ml of Bovine albumin primary standard and 1 ml of Biuret reagent were added into standard tube, 1 ml of Biuret reagent was added into blank tube.

• 3- The contents were mixed well, incubated at 37°C for 5min, standard and blank tubes.

• 4- The absorbance read at 540 nm.

• 5- Blank was used for zero adjustment of standard, and the test.

• A sample • Concentration of protein ( g/dL) = x 7.0 • Where: A standard • A: absorbance of recorded sample• 7.0: Standard concentration

Calculation:

• Adults: 6.6-8.7 g/dL (66-87g/L).• Prematures: 3.6-6.0 g/dL (36-60 g/L).• Newborns : 5.3-8.9 g/dL (53-89 g/L)• Pregnancy: Concentration lowers from 69 to

61 g/L

Reference values