Electrophoresis (2)

1. Seminar report on Electrophoresis
Topics:
•Moving boundary electrophoresis
•zonal electrophoresis
Date:9/1/14
Presented by: k.swapna
Guidence: Utham prasad SIR
Seminar report on Electrophoresis
Topics:
•Moving boundary electrophoresis
•zonal electrophoresis
Date:9/1/14
Presented by: k.swapna
Guidence: Utham prasad SIR

2. Electrophoresis
• Separation according to migration of charged
particles in electric field
• Electrolysis: Chemical decomposition
supplemented by current (components reach
electrode)
• F = v f = q E
V=Velocity, E=Electric field, q=Charge
f=Frictional coefficient

3. Electrophoresis
• Electrophoresis is a method whereby charged molecules
in solution, chiefly proteins and nucleic acids, migrate in
response to an electrical field.
• Their rate of migration through the electrical field,
depends on the strength of the field, on the net
charge, size, and shape of the molecules, and also on
the ionic strength, viscosity, and temperature of the
medium in which the molecules are moving.
• As an analytical tool, electrophoresis is simple, rapid
and highly sensitive.
• It can be used analytically to study the properties of a
single charged species or mixtures of molecules. It can
also be used preparatively as a separating technique

4. In most electrophoresis units, the gel is mounted between two
buffer chambers containing separate electrodes so that the
only electrical connection between the two chambers is through
the gel.

5. The Technique

6. Electrophoresis

7. Temperature and Electrophoresis
• Important at every stage of
electrophoresis
– During Polymerization
• Exothermic Reaction
• Gel irregularities
• Pore size
– During Electrophoresis
• Denaturation of proteins
• Smile effect
• Temperature Regulation of Buffers

8. Electrophoretic mobility
• Electrophoretic mobility μ = v / E = q / f
– V=Velocity, E=Electric field, q=Charge
f=Frictional coefficient
• Depends on
– Particle property :Surface, charge, density &
size
– Solution properties: Ionic strength, pH,
Conductivity,viscosity
– Temperature & Voltage

9. Types of electrophoresis-
Moving boundary
Electrophoresis

10. Moving boundary
Electrophoresis
• U shaped
• Multiple moving boundaries due to
different components
• Local changes in refractive index measured

11. Zonal Electrophoresis

12. Zonal electrophoresis
• Complete separation achieved
• Discrete zones formed
• 3 ways to stabilize components
– (i)Use of support material:
– Filter paper
– Membrane
– Gel

13. Zonal electrophoresis
– (ii)Density gradient stabilization:
- Sucrose,glycerol,ethylene glycerol
- Prevents convection current
(iii)Free zone electrophoresis
• Rotation of vessel during run
• Continuous film of buffer
perpendicular to electric field

14. Reference:
o Text book of Authour stream.

15. Thank you