Electrophoresis is an analytical technique that separates and analyzes ionized analytes using their migration in an electric field. Charged molecules are separated as they migrate through a porous medium like agarose or polyacrylamide gel under the influence of an electric field generated between electrodes. The direction of migration depends on the charge, with positively charged ions or DNA fragments migrating toward the negative cathode and negatively charged molecules toward the positive anode. Different electrophoresis methods like agarose gel electrophoresis and polyacrylamide gel electrophoresis are used depending on the size of DNA fragments to be separated.

1. 1
Clinical Laboratory Principle
Electrophoresis

2. Electrophoresis
• Analytical Technique
• The most commonly used in clinical applications.
• Separate and analyzed Ionize analytes
• Charged molecules migrate in a porous supporting
medium (agarose), after the sample is mixed with a
buffer solution, generates an electropherogram

3. Electrophoresis
Definition:
“ The migration of charged solutes or particles in a
liquid medium under the influence of an electrical
field.”

4. Theory of Electrophoresis
In an electrophoresis system, chemical take on electrical
charge and becomes ionized.
• Move toward electrode
Negative Electrode :Cathode
Positive Electrode : Anode
Positive ions(cations) migrate toward the Cathode
Negative ions (anions) migrate toward the Anode

5. Theory of Electrophoresis
CathodeAnode

6. Electrophoresis Apparatus
1. Buffer Tanks
2. Electrodes (Anode/Cathode)
3. Electrophoretic support for separation
4. Wick
5. Tank Cover
6. Power Supply

7. Electrophoresis Apparatus

8. Electrophoresis
 Agarose Gel Electrophoresis (AGE)
 Polyacrylamide Gels Electrophoresis (PAGE)

9. Agarose Gel Electrophoresis (AGE)
• Smaller DNA fragments have
migration rates in agarose that are
inversely proportional to the logs of
their molecular weights
• This relationship decreases as their
fragment size increases.

10. Loading of PCR Amplified Product in Agarose Gel

11. Direction
VerticalHorizontal

12. Agarose Gel Electrophoresis (AGE)

13. Agarose Gel Electrophoresis (AGE)

14. Visualization of PCR Amplified DNA Products

15. Polyacrylamide Gels (PAGE)
• Polyacrylamide is a polymer
• Prepared by heating acrylamide with a variety of catalysts,
with or without cross-linking agents.
• Polyacrylamide gel is
1. Thermostable
2. Transparent
3. Durable
4. Chemically Inert (uncharged)

16. Polyacrylamide Gels
• Capable of resolving DNA molecules that differ by as
little as 2% in length (1 bp in 50 bp).
• Accommodates a larger amount of sample (up to 10 fig) in
a single sample slot (well)
• DNA recovered from a polyacrylamide gel is extremely
pure, containing no inhibitors.
• Polyacrylamide is most useful for mixtures of smaller
DNA fragments and resolves fragments < 1 kbp

17. Apparatus for Polyacrylamide Gels (PAGE)