Serum protein electrophoresis is used to separate serum proteins into fractions based on their size and electrical charge. It has five major fractions: albumin, alpha1, alpha2, beta, and gamma globulins. Specific diseases and conditions cause abnormal patterns in the fractions. For example, liver disease decreases albumin levels while nephrotic syndrome increases alpha2 levels due to protein loss in the urine. The test is useful for identifying inflammatory states, immunodeficiencies, and monoclonal gammopathies.

1. NARENDRA YADAV
PAPER 4
SERUM ELECTROPHORESIS

2. INTRODUCTION
 Serum is the component that is neither a blood cell nor a
clotting factor; it is the blood plasma not including the
fibrinogens.
 Serum includes all the electrolytes, antibodies, antigens,
hormones, and any exogenous substances (e.g., drugs
and microorganisms).
 Serum is used in numerous diagnostic tests, as well as
blood typing.

3. COMPONENT OF SERUM
 Serum contains two major protein groups: albumin and
globulin.
 Using protein electrophoresis, these two groups can be
separated into five smaller groups (fractions):
1. albumin fraction
2. alpha 1 fraction
3. alpha 2 fraction
4. beta fraction
5.gamma fraction.

4. ALBUMIN
 Produce by liver.
 Largest protein component of serum.
 Moves toward anode durning electrophoresis.
 Usually a single band is seen.
 Heterozygous individuals may produce bisalbuminemia

5. ALPHA 1 FRACTION &
ALPHA 2 FRACTION
 Alpha 1 fraction component:
1.anti-trypsin=protease inhibitor.
2.thyroid binding protein= binding to thyroid hormones.
3.transcortin=Transcortin, transporting cortisol hormones.
 Alpha 2 fraction component:
1.ceruloplamin=copper-carrying protein.
2. haptoglobins= binds free hemoglobin released with
high affinity and thereby inhibits its oxidative activity.

6. BETA AND GAMMA FRACTION
 Beta fractions consist of:
1. transferrin=iron-binding glycoproteins.
2.lipoproteins= LDL& HDL.
 Gamma fraction cosist of:
 1. immunoglobulins.

7. ELECTROPHORESIS
 Electrophoresis is a laboratory technique in which the
sample is placed into a gel, and exposed to an electrical
current to separate the sample on basis of their charge or
size of the sample.
 Serum protein electrophoresis is a laboratory test that
examines specific proteins in the blood called globulins.
 Serum electrophoresis is used separate the serum protein
components into five major fractions by size and electrical
charge: i.e. serum albumin, alpha-1 globulins, alpha-2
globulins, beta globulins, and gamma globulins.

8. PRINCIPLE
 A charged particle placed in an electrical field migrates towards
the anode or cathode depending on the net charge carried by the
particle.
 Serum proteins carry a negative charge at pH 8.6 and move
toward the anode.
 Serum is placed into a sample within low EEO agarose gel in an
alkaline buffer.
 A standardized voltage is applied and upon staining, five distinct
protein bands (albumin, α1, α2, β, and γ) are seen.
 The membrane is then run through a densitometer and the light
absorbance by the protein bands is recorded.
 The absorbance of light is proportional to the relative protein
concentration. The protein concentration of each band is then
calculated.

9. PROTOCOL
 Preparation of agarose gel.
 Sample preparations.
 Electrophoresis.
 Staining.
 Destaining.
 Visualizing.

10. INTERPRETATION
 The densest band are of albumin.
Which run towards the anode durning
the electrophoresis.
 Alpha 1 fraction
 Alpha 2 fraction
 Beta fraction
 Gamma fraction which move towards
the cathode durning the
electrophoresis. The band is
observed back of the origin.

11. Normal Graph after interpretation

12. PATHOLOGICAL CONDITION ASSOCIATED WITH A CHARACTERISTIC
SERUM PROTEIN ELECTROPHORESIS PATTERN.

13. ALBUMIN DISORDER
(HEPATIC CIRRHOSIS)
 Abnormal function of
liver.
 Protein synthesis is
compromised and the
concentration of albumin
and protein in the alpha
and band are decreased.
 Additional common
finding is beta-gamma
bridging due to
increased IgA.

14. NEPHROTIC SYNDROME
 Nephrotic syndrome is a
kidney disorder.
 Loss of protein through the
kidneys(proteinuria) leads
to low protein levels in the
blood.
 Synthesis cause its
accumulation.
 Large proteinuria is due to
an increase in permeability
of the "filtering membrane"
of the kidney, which
selectively filter only
albumin and other smaller
molecular.
 Alpha2 is sufficiently large
so that it is not filtered and
increased Synthesis cause
its accumulation.

15. ALPHA 1
ANTITRYPSIN -DEFICIENCY
 Alpha 1-antitrypsin
deficiency (α1-antitrypsin
deficiency, A1AD) is a
genetic disorder that
causes defective production
of alpha 1-antitrypsin
(A1AT), leading to
decreased A1AT activity in
the blood and lungs.

16. ALPHA1 & ALPHA2 DISORDER
ACUTE INFLAMATION
 There is ncreased alpha 1
and alpha2 band durning the
inflammatory response from
increased hepatic synthesis
of acute phase reactant
protein.

17. GAMA FRACTION DISORDER
CHRONIC INFLAMATION
 Immunoglobulin synthesis
by the activated B
lymphocytes transformed to
plasma cells is
demonstrated by the
increased polyclonal gamma
band.

18. IMMUNOGLOBULIN DEFICIENCY
 Deficient
immunoglobulins
synthesis is
demonstrated by
deminsihed gamma
band.

19. MONOCLONAL GAMMOPATHY
 Usually sharp band in the
gamma region suggest the
presence of a homogenous
immunoglobulins.
 Homogenous
immunoglobulins are also
found in waldenstrom’s
macroglobulinemia

20. Serum Protein Fraction Increased Decreased
Albumin Severe dehydration
Malnutrition, cachexia, liver
disease, nephrotic syndrome,
protein-losing enteropathies,
severe burns
Alpha-1 Inflammatory states, pregnancy Alpha-1 antitrypsin deficiency
Alpha-2
Inflammatory states, nephrotic
syndrome, oral contraceptive use,
steroid use, hyperthyroidism
Hemolysis, liver disease
Beta
Hyperlipidemia, iron-deficiency
anemia
Hypo-B-lipoproteinemia,
malnutrition
Gamma
Polyclonal and Monoclonal
Gammopathies
Agammaglobulinemia,
hypogammaglobulinemia
Various disease states or conditions alter the pattern of proteins in electrophoresis

21. “
”
Thank you

22. “
”
REFERENCE
GOOGLE
WIKIPEDIA
BIOTECHNOLOGY JOURNAL