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Sample

Indication

Pathophysiology

  1. Definition of electrophoresis: This is the migration of charged solutes or particles in a liquid medium under the influence of the electrical field.
  2. Analytes of interest are:
    1. Proteins.
    2. Peptides.
    3. Aminoacids.
    4. Nucleic acid.
    5. OLigonucleotides.
    6. Cations in body fluids and the tissue.
  3. Proteins are constituents of muscles, hormones, hemoglobin, and transport protein,
    1. Serum proteins are a source of nutrition and buffer system.
    2. Immunoglobulins have a function in the immune system.
  4. Carrier proteins like haptoglobins, prealbumin, and transferrin, transport certain ions, and molecule to their site of action.
  5. Some of the protein regulates:
    1. The activity of various proteolytic enzymes.
    2. The osmotic pressure within the vascular compartment.
    3. Metabolic substances like a hormone.
  6. Immunoglobulins are a major component of gamma-globulins.
    1. In the normal person, the immunoglobulins are of polyclonal nature.
    2. Monoclonal band of immunoglobulin indicates a neoplastic process like multiple myeloma and Waldenstrom's macroglobulinemia.
  7. Serum electrophoresis separate proteins into 5 different bands:
    1. Albumin, and Prealbumin. Albumin is formed in the liver and is 60% of total proteins.
    2. alpha1-globulin (α1-globulin).
    3. alpha 2-globulin (α2-globulin).
    4. beta – globulin (β-globulin).
    5. gamma – globulin (γ-globulin).
Protein Adult value g/dL Cord blood g/dL Mother's serum g/dL
Albumin 3.5 to 5.0 3.3 4.2
α1-Globulin o.1 to 0.4 0.0 0.3
α2-Globulin 0.3 to 0.8 0.4 1.2
β-Globulin 0.6 to 1.1 0.7 1.3
γ-Globulin 0.5 to 1.7 1.0 1.3

  1. These different bands have a different zone with the characteristic presence of different proteins. e.g.
    1. Albumin zone shows the only albumin.
    2. alpha1- zone shows :
      1. alpha1 – lipoprotein.
      2. High-density lipoprotein.
      3. alpha 1 -antitrypsin.
    3. alpha 2 zone shows :
      1. alpha 2 macroglobulins.
      2. Haptoglobin.
      3. β -lipoprotein.
    4. beta zone shows :
      1. Transferrin.
      2. Complement 3 (C3).
    5. gamma zone shows :
      1. Fibrinogen.
      2. IgA.
      3. IgM.
      4. IgG.

Functions of various proteins:

  1. Albumin :
    1. maintain the colloidal osmotic pressure.
    2. Albumin also transports drugs, hormones, and enzymes.
    3. Albumin measures the liver function. In liver diseases, its concentration will be markedly low when there is liver cell damage.
  2. The half-life of Albumin is 12 to 18 days, so the liver cell damage will be shown after this period.
  3. Globulins :
    1. These are the main component of immunoglobulins (Antibodies).
    2. Some are transporting proteins like thyroid and cortisol binding protein.
    3. Haptoglobin binds hemoglobin during hemolysis.
    4. Ceruloplasmin is a carrier of copper.
    5. They may also act as a transport vehicle.

Principle of Electrophoresis

  1. This is basically separation or migration of charged solutes of particles in liquid medium under the influence of the electrical field.
  2. Chemical substances carrying charges because of ionization moves towards either cathode ( Negative electrode ) or anode ( positive electrode ). So the protein ions ( cation ) moves towards the cathode, and negative ions ( anion ) moves towards the anode.
  3. Serum Electrophoresis can separate the various components of blood proteins into bands or zones according to their electrical charge under the influence of electrical current.
    1. Zones of proteins are separated from the neighboring zones.
    2. Then these zones are visualized by the stains.
    3. The support medium is dried and can quantify each zone by the densitometer.
    4. The support medium can be kept permanently after drying.
  4. The rate of migration depends upon:
    1. The net electrical charge of the molecule.
    2. Size and shape of the molecule.
    3. Electrical field strength.
    4. The temperature of the medium.
    5. Properties of the support medium.
  5. Urine electrophoresis classifies renal protein-losing nephropathies.
  6. CSF electrophoresis diagnose monoclonal band.

Components needed for electrophoresis are:

  1. Tank with power supply (electrical field).
  2. Electrode, either carbon or platinum.
  3. Buffer system (in the buffer tanks).
  4. support media like :
    1. agar gel.
    2. agarose.
    3. Cellulose acetate.
    4. Polyacrylamide.
  5. Stains. This is used to visualize the separate bands.
  6. Densitometer to read the different bands and quantitate them.
  7. The automated system does automatically electrophoresis instead of a manual method.

Normal

Source 1

Total Proteins in Adult

 Age  g/dL
 Cord blood 4.8 to 8.0 
 Premature 3.6 to 6.0 
 Newborn 4.6 to 7.0 
 One week  4.4 to 7.6 
7 months to one year 5.1 to 7.3
 1 to 2 years  5.6 to 7.5
≥3 years 6.0 to 8.0
Adult  
Ambulatory 6.4 to 8.3
Recumbent 6.0 to 7.8
>60 years Lower by ∼0.2 

Source 2

Children

Source 1

Protein electrophoresis on cellulose acetate:

Age g/dL
Albumin
Adult 3.5 to 5.0
α1- Globulin
Adult 0.1 to 0.3
α2- Globulin 
Adult 0.6 to 1.0
High in children <15 years  
β- Globulin 
Adult  0.7 to 1.1
γ-Globulin 
Adult 0.8 to 1.6

Protein electrophoresis on Agarose:

Age g/dL
Albumin
0 to 15 days 3.0 to 3.9
15 days to one year 2.2 to 4.8
1 to 16 years 3.6 to 5.2
>16 years 3.9 to 5.1
α1- Globulin
<1 year 0.1 to 1.03
1 to 16 years 0.1 to 0.4
>16 years 0.2 to 0,4
α2- Globulin
0 to 15 days 0.3 to 0.6
15 days to one year 0.5 to 0.9
1 to 16 years 0.5 to 1.2
>16 years 0.4 to 0.8
β- Globulin
0 to 15 days 0.4 to 0.6
15 days to one year 0.5 to 0.9
1 to 16 years 0.5 to 1.1
>16 years 0.5 to 1.0
γ-Globulin
0 to 15 days 0.7 to 1.4
15 days to one year 0.5 to 1.3
1 to 16 years 0.5 to 1.7
>16 years 0.6 to 1.2

Increased Total Protein (Hyperproteinemia) seen in:

  1. Dehydration and hemoconcentration due to a fluid loss like vomiting, diarrhea, and poor kidney function.
  2. Liver diseases.
  3. Multiple myelomas.
  4. Gammopathies.
  5. Waldenstrom’s globulinemia.
  6. Sarcoidosis and other granulomatous diseases.
  7. Autoimmune diseases like SLE, and Rheumatoid arthritis.
  8. Chronic inflammation.

Decreased Total protein (Hypoproteinemia) seen in:

  1. Severe liver disease.
  2. Renal disease, nephrotic syndrome.
  3. Starvation and malabsorption.
  4. Diarrhea, ulcerative colitis, and Crohn’s disease.
  5. Severe burn.
  6. Skin diseases.
  7. Hypothyroidism.
  8. Heart failure.

Increased Albumin is seen in:

  1. Dehydration.
  2. Intravenous infusion.

Decreased Albumin is seen in:

  1. Liver diseases.
  2. Alcoholism.
  3. Malabsorption.
  4. Nephrotic syndrome.
  5. Starvation.
  6. Protein-losing enteropathy like Crohn’s disease and Whipple's disease.
  7. Ascites.
  8. Congenital albuminemia.
  9. Pregnancy.
  10. Overhydration.
  11. Autoimmune diseases like systemic lupus erythematosus.

Increased α-1 globulin seen in:

  1. Biliary Cirrhosis.
  2. Obstructive jaundice.
  3. Multiple myelomas.
  4. Kidney disease like nephrosis.
  5. Acute and chronic infection.
  6. Ulcerative colitis.

Decreased α-1 globulin seen in:

  1. Kidney diseases like nephrosis.
  2. Acute Hemolytic anemia.
  3. Juvenile pulmonary emphysema.

Increased α-2 globulin seen in:

  1. Kidney disease like nephrotic syndrome.
  2. The inflammatory disease that is due to an increase in the acute phase proteins.

Decreased α-2 globulin seen in:

  1. In hemolysis because haptoglobin is alpha 2 globulin which will decrease in hemolysis.
  2. Wilson’s disease.
  3. Hyperthyroidism.
  4. The liver disease when liver function is defective.

Increased β- globulin is seen in:

  1. Biliary Cirrhosis.
  2. Obstructive jaundice.
  3. Neoplasm like multiple myeloma.
  4. In all hypercholesterolemia which may be due to hypothyroidism and nephrosis.
  5. iron deficiency anemia.

Decreased β- globulin is seen in:

  1. Kidney diseases like nephrosis.
  2. malnutrition.

Increased  γ- globulin is seen in:

  1. Hepatic diseases like cirrhosis.
  2. acute and Chronic infections.
  3. Multiple myelomas.
  4. Autoimmune diseases like SLE and rheumatoid arthritis.
  5. Waldenstrom’s macroglobulinemia.
  6. Leukemia.
  7. Malignancies like Hodgkin's lymphoma, lymphomas,

Decreased γ- globulin is seen in:

  1. Nephrotic syndrome.
  2. Agammaglobulinemia.
  3. hypogammaglobulinemia.
  4. An immune deficiency which may be due to infections, steroid therapy, or lymphomas.

Monoclonal Gammopathy 

                                                           


Possible References Used

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