Depending on the kind of charge they carry, iortized solutes move toward either tbe cathode (negative electrode) or the anode (positive electrode) in an electrophoresis system. For example, positive ions (cations) migrate to the cathode and negative ions (anions) to the anode. An ampholyte (a molecule that is either positively or negatively charged, formerly called a zwitterion) becomes positively charged in a solution more acidic than its isoelectric point (pI)* and migrates toward the cathode. In a more alkaline solution) the ampholyte becomes negatively charged and migrates toward the anode. Because proteins contain many ionizable amino (-NH,) and carboxyl (-COOH) groups, and the bases in nucleic acids may also be positively or negatively charged, they both behave as ampholytes in solution.
Rate of migration of ions depends on:
1. Net Charge of molecule
2. Size and shape
3. Strength of electric field
4. Properties of the supporting medium
5. Temperature
Driving Force(F):
Resisting force(F’):
These two counteracting force produces a constant velocity
The last equation gives Electrophoretic mobility(µ)
Electrophoretic mobility is the rate of migration (V) per unit field strength (X)
µ= cm^2/(V)(sec)
- Is directly proportional to net charge
Inversely proportional to
- size of molecule
- Viscosity of medium
Another factor and potential problem that can affect mobility is wick flow. During electrophoresis heat evolved because of the passage of current through a resistive medium can cause evaporation of solvent from the electrophoretic support. This drying effect draws buffer into the support from both buffer compartments. If significant, the flow of buffer from both directions can affect protein migration and hence the calculated mobility.
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