AAT Bioquest

How Does Protein Purification Work?

Posted March 16, 2022


Protein purification is a process that aims to isolate individual proteins and protein complexes of interest in order to analyze their interactions with other proteins, DNA, or RNA. While there are different types of protein purification strategies, all involve 4 basic steps:

  • Cell lysis – In this first step, cells are disrupted and proteins denatured using various chemical, enzymatic, or physical methods. 
  • Clarification – In the second step, filtration or centrifugation are used to remove all contaminants and obtain only the proteins to be purified. The protein extract is now ready for concentration.
  • Protein Binding – The goal of this step is to isolate the protein from the extract. Techniques such as affinity chromatography and magnetic bead separation are used to isolate the protein by binding the protein molecule to a matrix. Protein binding is also called the recovery stage.
  • Elution – In this last step, an elution buffer is used to extract one material that is adsorbed to another and eliminate all the non-specific binding on the column. The number and duration of the washes vary depending on the type of column used.

Completion of these 4 protein purification steps helps researchers obtain high protein concentrations that can then be used for further analysis.

Additional resources

Protein production and purification

ProLite™ His-Tag Protein Gel Staining Kit *Green Fluorescence*