Biochemistry Determination of Total Protein in Serum PDF

Almaaqal University Biochemistry Determination of Total Protein in Serum Dr/ Wael Sobhy Darwish Biochemistry PhD  Total protein The proteins are organic...

Almaaqal University Biochemistry Determination of Total Protein in Serum Dr/ Wael Sobhy Darwish Biochemistry PhD  Total protein The proteins are organic macromolecular compounds, widely distributed in the organism. Proteins are important for body growth and health. Plasma proteins are synthesized mainly by the liver with the exception of γ – globulins which are synthesized by plasma cells present in lymph nodes, spleen and bone marrow. The total protein test measures the concentration of all the proteins in the plasma of your blood (with the exception of fibrinogen as the analysis is on a clotted blood sample). Total protein measures the amount of two types of proteins, albumin and globulin. Function of plasma protein Transport (Albumin, prealbumin, globulins) Maintain plasma osmotic pressure (Albumin) Defense (Immunoglobulins and complement) Clotting and fibrinolysis (Thrombin and plasmin)  How is it used? Total protein measurements can reflect someone's nutritional status. The presence of kidney disease and liver disease, as well as many other conditions. If total protein is abnormal, further tests may be needed to identify which protein fraction is abnormal, so that a specific diagnosis can be made.  When is it requested? Total protein is measured along with several other tests to provide information if you have symptoms that suggest a liver or kidney disorder, or to investigate the cause of abnormal pooling of fluid in tissue (oedema). It will also be requested as part of the investigation of multiple myeloma.  ASSAY PRINCIPLE The reaction is based on the conversion of nitrogen in proteins and other compounds to ammonia This method is based on the biuret reaction where divalent copper reacts with the peptide bonds of protein under alkaline conditions to form the characteristic pink to purple biuret complex.  PROCEDURE 1. Assay conditions: Wavelength: 530 - 570 nm Cuvette: 1 cm light path Temperature: 37ºC /15-25ºC Sample Serum with out any preparation  PROCEDURE Blank Standard Sample WR(mL) 1.0 1.0 1.0 Standard (µL) -- 20 -- Sample (µL) -- -- 20 Mix and incubate for 5 min. at 37ºC or 10 min. at room temperature Read the absorbance (A) of the samples and Standard, against the Blank. The colour is stable for at least 60 minutes  Calculation Total Protein concentration = Expected Value Adult: 6.3 – 8.3 g/dl Child : 6.0 – 8.0 g/dl What does the result mean? A- Hypoproteinemia: 1- Prolonged starvation. 2- Malnutrition and malabsorption such as in Celiac disease. 3- Decreased synthesis by the liver such as in liver cirrhosis or failure. 4- Increased loss of proteins as in kidney disease (e.g., nephrotic syndrome) 5. Excessive bleeding. 6. Extensive burns.  B- Hyperproteinemia: 1- Increased synthesis such as in infections like viral hepatitis B or C, or HIV. 2- Increased synthesis in some cancers such as multiple myeloma or certain types of lymphoma.  3- Dehydration that results from conditions like vomiting, diarrhea, excessive sweating, and diabetic acidosis  If there any thing else I should know Prolonged application of a tourniquet or standing during blood collection can increase total protein levels. Many medications may affect total protein levels, including oestrogens, steroids, and oral contraceptives.