The FLAG tag (peptide sequence DYKDDDDK) is a short, hydrophilic protein tag commonly used in conjunction with antibodies in protein pull-downs to study protein–protein interactions. The FLAG tag may be inserted at the N terminus, the N terminus preceded by a methionine residue, the C terminus, or internal positions of the target protein. Because of its hydrophilic nature, the FLAG tag is commonly found on the surface of a fusion protein, which makes it more available as an epitope for binding to antibodies. In addition, the high hydrophilicity and small size of the FLAG tag tend to interfere less with protein expression, proteolytic maturation, antigenicity, and function. FLAG tags are also easily removed by enterokinase (EK). We offer quality antibodies specific to FLAG tag that can be used in a variety of research needs. Fluorescent and enzyme-conjugated versions of our FLAG Tag antibodies are available for your added convenience.

All antibodies listed here recognize a similar epitope to the ANTI-FLAG® M2 antibody of Sigma-Aldrich. FLAG is a registered trademark of Merck KGaA, Darmstadt, Germany and ANTI-FLAG is a trademark of Sigma-Aldrich Co. LLC.

See all Anti-DYKDDDDK tag antibodies (bind to the FLAG sequence)
 

Applications
Immunofluorescence analysis of FLAG Tag using anti-DYKDDDDK recombinant polyclonal antibody

For immunofluorescence analysis DYKDDDDK—BNIP3 transfected HEK293 cells were fixed and permeabilized for detection of DYKDDDDK-Tag protein using DYKDDDDK Tag Recombinant Polyclonal Antibody (Cat. No. 740001,
1 µg/mL) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal Recombinant Secondary Antibody, Alexa Fluor 488 (Cat. No. A27034, 1:2000).Panel a) shows representative cells that were stained for detection and localization of DYKDDDDK-Tag protein (green), Panel b) is stained for nuclei (blue) using SlowFade Gold Antifade Mountant with DAPI (Cat. No. S36938).Panel c) represents cytoskeletal F-actin staining using Rhodamine Phalloidin (Cat. No. R415, 1:300).Panel d) is a composite image of Panels a, b and c clearly demonstrating cytoplasmic localization of DYKDDDDK - BNIP3. Panel e) shows no signal with untransfected cell and panel f) represents control cells with no primary antibody to assess background.

Western blot analysis of FLAG Epitope Tag was performed by loading various amounts of E. coli lysate containing a multi-epitope tagged protein per well onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a low fluorescence PVDF membrane and blocked with Fish Serum blocking buffer for at least 1 hour. The membrane was probed with DYKDDDDK Tag Monoclonal Antibody (L5), Alexa Fluor 488 (Cat. No. MA1-142-A488) at a dilution of 1:500 for 1 hour at room temperature on a rocking platform and washed in TBS-0.1% Tween-20. Detection was performed using a fluorescence imaging system.

Western blot analysis of FLAG epitope tag using FLAG Tag Alexa Fluor 488 monoclonal antibody

Resources

For Research Use Only. Not for use in diagnostic procedures.

For Research Use Only. Not for use in diagnostic procedures.