Protein Separation Techniques

Isoelectric Point

The isoelectric point is the pH at which a molecule is neutral. If a protein or amino acid possesses many acidic side chains, its isoelectric point will be low. Conversely, if a protein or amino acid possesses many basic side chains, its isoelectric point will be high. This characteristic can be used to separate proteins through a process called electrophoresis.

Electrophoresis

Electrophoresis takes advantage of varying isoelectric points to separate proteins.

  1. A pH gradient is set up on a gel matrix
  2. An electric current is passed through the gel to create an anode and cathode.
  3. Proteins which are in a region below is isoelectric point (low pH) will be positive and migrate to the cathode until it reaches its isoelectric point.
  4. Proteins which are in a region above its isoelectric point (high pH) will be negative and migrate to the anode until it reaches its isoelectric point.

 

The following figure demonstrates the process of electrophoresis:

Gel Electrophoresis
By Mrbean427 (Own work) [CC BY-SA 3.0 (http://creativecommons.org/licenses/by-sa/3.0) or GFDL (http://www.gnu.org/copyleft/fdl.html)], via Wikimedia Commons

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