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

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

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

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.

The Technique

Electrophoresis

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

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

Types of electrophoresis- Moving boundary Electrophoresis

Moving boundary Electrophoresis

• U shaped

• Multiple moving boundaries due to different components

• Local changes in refractive index measured

Zonal Electrophoresis

Zonal electrophoresis

• Complete separation achieved

• Discrete zones formed

• 3 ways to stabilize components– (i)Use of support material: – Filter paper– Membrane– Gel

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

Reference:o Text book of Authour stream.

Thank you