THE DIAGNOSTIC USES OF ELECTROPHORESIS

CPY 703 SEMINAR PRESENTATION BY

DAMILOLA SAMSON AKINDUKO MATRIC No: 160388

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PRESENTATION OUTLINE
ELECTROPHORESIS

-Introduction -Principle -Set-up CLINICAL APPLICATIONS

GENERAL PROCEDURE IN ELECTROPHORESIS ELECTROPHORETIC PATTERNS IN DISEASES

CONCLUSION
REFERENCES
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ELECTROPHORESIS
INTRODUCTION Electrophoresis is a technique for separation of different charged particles.
It is based on movement of charged particles through a solution

when subjected to an electrical field.

PRINCIPLE In an electrophoresis system, particles take on electrical charge, move toward either the cathode(negative electrode) or the anode(positive electrode).
Separation is based on the differences in their charge-to3

mass ratios as well as their molecular weights.

CONVENTIONAL ELECTROPHORESIS

SET-UP

(Adapted from Karcher & Landers, 2006)

1. 2. 3.
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4.

Two buffer boxes containing the buffer solution Electrodes connected to the power supply. The electrophoresis support which contacts the buffer by means of the wicks. Wicks

BUFFERS The buffer carries the applied current and also determines the electrical charge on the particles.
SUPPORT MEDIA The support medium provides the matrix on which separation takes place. Types of support media include: starch, cellulose acetate, agarose and polyacrylamide gels. CLINICAL APPLICATIONS To extensively study families of proteins in order to detect 5 genetic- or disease-based differences;

CLINICAL APPLICATIONS (CONTINUED)

For diagnosis and monitoring of multiple myeloma; Detection of large structural alterations in DNA

which cause mutation, e.g. deletions, insertions, duplications, etc.;

To detect in urine the presence of immunoglobulin

side chains (Bence Jones protein); etc.

GENERAL PROCEDURES IN ELECTROPHORESIS

SEPARATION
The support medium (agarose or polyacrylamide gel) placed

in the electrophoresis chamber;

The sample is then added to the support and made to contact

the buffer indirectly by means of the wicks or tissue paper;

Electrophoresis is then conducted for a determined length of

time once the current is applied. STAINING

The support medium is stained using dyes(e.g. Amido Black B
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or Coomassie Brilliant Blue);

DETECTION AND QUANTIFICATION The support medium is placed in a densitometer in which the gel (or other support medium) is moved past a measuring optical system;
The absorbance of each fraction is displayed on a

recorder chart or an electronic display.

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(Adapted from Crook, 2006)

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(Johnson, 2005)

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(Johnson, 2005)

PATHOLOGIC SERUM PROTEIN ELECTEOPHORESIS PATTERNS

o THE ACUTE PHASE PATTERN

Synthesis of the acute-phase proteins is a direct result inflammation triggered by tissue damage of any kind.

o CHRONIC INFLAMMATORY STATES A diffuse rise in -globulin coupled with the usual increase in the -1 and -2 fractions of the acute phase. 13

PATHOLOGIC SERUM PROTEIN ELECTEOPHORESIS PATTERNS(continued)

o CIRRHOSIS OF THE LIVER:

Albumin and alpha1-globulin: reduced, -globulin: markedly increased, with apparent fusion of the - and bands. o NEPHROTIC SYNDROME Reduced albumin, alpha1 and sometimes -globulin; Increase in alpha2-globulin due to a relative increase in 2macroglobulin.
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PATHOLOGIC SERUM PROTEIN ELECTEOPHORESIS PATTERNS(continued)

o 1-ANTITRYPSIN(AAT)

DEFICIENCY:

The 1-band consists almost entirely of AAT : absence or an obvious reduction in the density strongly suggests AAT deficiency.

o HYPOGAMMAGLOBULINEMIA Decrease in plasma immunoglobulins; Total protein concentration 15 usually low.

PATHOLOGIC SERUM PROTEIN ELECTEOPHORESIS PATTERNS(continued)

o MONOCLONAL GAMMOPATHY Presence of excessive amounts of a single monoclonal immunoglobulin.

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CONCLUSION
It is recommended that full use of this diagnostic technique be made for better diagnostic sensitivity and

specificity.
In addition, correct monitoring of buffer pH,

voltage, equal
quantity of test and control applied and skill of
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REFERENCES
Berg JM, Tymoczko JL Stryer L (2002).

Biochemistry (5th ed.). WH Freeman. ISBN 07167-4955-6. http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=stry er.section.438#455.

Crook MA. Proteins in plasma and urine. Clinical

Chemistry and Metabolic Medicine, 7th ed. London: Edward Arnold, 2006:280-300.
Johnson AM. Amino acids, peptides and proteins.
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In: Burtis CA, Ashwood ER, Bruns DE, eds. Tietz textbook of clinical chemistry and molecular

REFERENCES (CONTINUED)
Karcher RE, Landers JP. Electrophoresis. In:

Burtis CA, Ashwood ER, Bruns DE, eds: Tietz textbook of clinical chemistry and molecular diagnostics, 4th ed. Philadelphia: Saunders, 2006:121-40.
Landers JP. Molecular diagnostics on

electrophoretic microchips. Anal Chem 2003;75:2919-27.

Viscaris P, Landers JP. Unconventional detection
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methods for microfluidic devices. Electrophoresis

THANK YOU ALL

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