Flag Tag Antibody

The Flag tag is a hydrophilic octapeptide (DYKDDDDK) with a molecular weight of approximately 1.0 kDa. It is commonly fused to the N- or C-terminus of proteins via recombinant DNA technology. Due to its small size and high hydrophilicity, the Flag tag typically does not interfere with protein expression, proteolytic maturation, or antigenicity. Moreover, the purification process for Flag-tagged proteins is non-denaturing, enabling the recovery of functionally active fusion proteins.

There are three commonly used monoclonal antibodies against the Flag tag: M1, M2, and M5.

M1 antibody recognizes the first four amino acids of the Flag sequence and requires a free N-terminal amino group, making it specific for N-terminal Flag-tagged proteins. It does not recognize Met-Flag fusion proteins.

M2 antibody binds independently of calcium ions (Ca^²⁺) and is suitable for both N-terminal (Met-Flag) and C-terminal Flag-tagged proteins.

M5 antibody shares similar properties with M2 but exhibits higher affinity for N-terminal Met-Flag fusion proteins.