Epitope tagging is a procedure that inserts a short amino acid sequence into a protein within an expression vector via genetic engineering. Antibodies that recognize the tag can then be used to detect the protein when no antibody to the target protein exists or when the target protein shows low immunogenicity. Such tags can be inserted at the C-terminus, N-terminus or even within the protein itself. Terminal insertion is preferred as internal tags may alter protein function or become buried within the tertiary structure of the protein. Initially tags were used to facilitate purification (e.g. 6xHis®, GST), however they are now used in Western blots, IHC, immunoprecipitation and flow cytometry.
One particularly powerful technique is using epitope tags in co-immunoprecipiation to study protein interactions. In this procedure, an epitope-tagged bait protein is initially transfected into cells. After cell lysis, the bait protein is immunoprecipitated with an anti-tag antibody-bead conjugate. Any proteins that interact with the bait protein will co-immunoprecipitate, and the resulting protein complex can then be analyzed by SDS-PAGE.
Two of the most popular commercial epitope tags are the FLAG® and OLLAS tags. The FLAG® tag (DYKDDDDK) contains the Enterokinase Cleavage Site (DDDK) which allows removal of the tag after purification. The OLLAS tag (E. coli OmpF Linker and Mouse Langerin fusion sequence) has been shown to be over 100 times more sensitive than other tags, including FLAG®,providing superior performance in all applications.
Novus not only offers antibodies to any of these tags, but also single and double tag cloning vectors for tagged protein expression. Please contact our technical support department (technical@novusbio.com) with any questions or for help choosing the optimal tag for your experiment.
