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Saturday, July 13, 2013

Steps in Protein Purification



  • Know properties of your Protein
  • Developing a Strategy for Purification
  • Choosing Suitable method for highest yield.
  • Optimize the Protein Purification
steps in protein purification CIPP

Know Your Protein

This is very important step to consider before developing the purification strategy. Knowing your protein means characterizing the protein. Understanding pI value, and other characteristics of protein will aid in the protein purification. Some of the techniques used for protein characterization are:

1. SDS- PAGE
2. Iso Electric Focussing
3. Western Blot
4. Mass Spec, etc

Developing a Strategy for Purification

Once the protein is characterized, it is easy to develop a strategy for purification. Depending on the stability and required purity suitable  method should be choosen. Keep in mind about the scale up during the strategy development. Some of the Common Protein purification mehod includes

  1. Protein Salting out using suitable salts
  2. Chromatographic Techniques like Ion Exchange Chromatography, Size Exclusion chromatography, Hydrophobic interaction Chromatography, Reverse Phase Chromatography, Affinity Chromatography, Etc.
  3. Protein Concentration through Filtration
  4. Lyophilization
Choosing Suitable method for highest yield.
For various reasons suitable method which provides highest yield need to be selected, a few to cite here is the production cost, one should keep in mind about the production cost involved while choosing a method for purification when it comes to scale up all these adds up so one should select a method which will give highest yield. chosen method used should not compromise the protein efficacy, activity, usablity and purity specifically when trying to purify a protein for therapeutic use.

Optimizing Protein Purification

Optimization is important step in any process not only just protein purification. Optimizing a process can have multiple advantages reduced time, improved yield, higher purity, more stability, cost efficient all these are some of the key target points to achieve by optimizing a process. Optimizing targets vary depending on the type of process and the final goal.