Detection and purification of proteins and peptides depend on specific antibodies. There are many commercially available antibodies however they do not cover all proteins, especially the novel proteins, proteins of unknown sequences and proteins or peptides of low antigenicity. Raising an antibody to a protein of interest can be quite time-consuming and expensive. A convenient solution for detection and purification of various proteins is offered by Epitope tags (also known as affinity or fusion tags). Epitope tagging is a common experimental technique by which a well characterized and specific sequence of amino acids called the “epitope” (recognized by a particular antibody) is combined with a protein of interest. Antibodies directed against these specific epitope tags can bind to the fusion proteins bearing the epitope, thus allowing the detection of these tagged proteins. These epitope tags allow detection and purification without disturbing the structure of the protein to which they are fused. Choice of tag depends upon many factors, and one of the most important considerations is the downstream analysis of the protein such as western blot analysis, immunoprecipitation, and affinity purification.
Advantages of Epitope Tagging
- Saves time as epitope tagging is quicker than a custom antibody preparation, which requires a few months.
- Single epitope tagging is also cheaper than production cost of an antigen specific antibody.
- Same epitope tag can be used for various proteins of different sizes.
- The epitope tag is composed of 3 to 14 amino acids and does not disrupt normal protein functions.
- The technique can be used to detect proteins that are very difficult to isolate and purify.
The epitope tagged proteins are commonly used for western blotting and immunoprecipitation. They can also be used for applications like protein purification, immunofluorescence microscopy and flow cytometry.