sERUM PROTIENS AND AG RATIO, TOTAL PROTIENS, ALBUMIN, GLOBULINS, TOTAL ALBUMIN, SERUM GEL ELECTROPHORESIS

1. SERUM PROTEINS & A/G
Ratio

2. • Plasma refers to the fluid portion of anticoagulant added
blood after cells are removed by centrifugation
• Serum is the fluid component of blood after it is allowed to
clot

4. Albumin
Most abundant plasma protein (~50%)
Synthesized by hepatocytes
Single polypeptide - 66 kDa
High no. of charged AA results in ↑ solubility
Plasma half life – 15 to 19 days
Acts as a transporter and major component of
colloid osmotic pressure (oncotic pressure)

5. Globulins
 Heterogeneous family of proteins
 Mol weight: 12-900 kDa
 Less water soluble than albumin
Types
Alpha (α) :
α1 (α1-antitrypsin, α1-acid glycoprotein, HDL)
α2 (Haptoglobin, α2-macroglobulin, Ceruloplasmin)
Beta (β) :
β1 (Transferrin,C4,LDL)
β2 (C3, β2-microglobulin)
Gamma (γ) : Immunoglobulins, C-reactive protein

6. Methods of Protein Estimation
Copper based
– Biuret assay (Biuret reagent)
– Lowry assay (Folin-Ciocaltau reagent)
– BCA assay (Bi-Cinchoninic Acid reagent)
Dye based
– Bradford assay

7. Principle
Compounds with 2 or more peptide bonds,
react with CuSO4 in alkaline conditions to
produce a complex which is violet to blue
colored.

8. Biuret Reaction

10. Lowry assay

11. BCA assay

12. Bradford assay

13. Bromo-Cresol Green Method
(Albumin analysis)
• BCG reagent at pH 4.3 is negatively charged
• pI of Albumin is 4.7
• Bromocresol green binds quantitatively with
albumin to from an intense bluegreen
• complex with an absorbance max at 670 nm.

17. A-G Ratio
Direct:
 Albumin (A) & Globulins (G) in the serum sample
are separated & determined separately
Indirect:
Serum total protein: Biuret reaction
Serum albumin: Bromocresol Green (BCG)
Serum globulin = Serum total protein - Serum
albumin

18. Hypoproteinemia
Hypoproteinemia is serum protein level < 6.7 g/dL
↓ (A/G)
↑ Loss
• Renal disorders(e.g. nephrotic syndrome)
↓ Synthesis
• Chronic liver disease(cirrhosis)
↑ (A/G)
• Due to decrease in globulin
• Agammaglobulinemia/
• hypogammaglobulinemia

19. Hyperproteinemia
Hyperproteinemia is serum protein level > 8.3 g/dL
Normal (A/G)
• Relative hyperproteinemia
• Dehydration
↓ (A/G)
• Due to increase in globulin
• Chronic infections
• Malignant proliferation of plasma cells -
e.g.myeloma

20. Serum protein electrophoresis

21. Serum Protein Electrophoresis

22. Liver damage Kidney damage