The original DYKDDDDK peptide (the same epitope recognized by Sigma's Anti-FLAG®** antibodies) was an 11-amino-acid leader peptide of the gene-10 product from bacteriophage T7 fused at the amino-terminus of GAL4 (yeast transcription factor). The binding reaction of this tag to the anti-DYKDDDDK antibody is calcium dependent, so proteins can frequently be eluted from an affinity matrix by buffers containing EDTA. It also allows for the tag to be placed at the amino-terminus, at the carboxy-terminus, or in association with other tags.
Buy one FLAG antibody, Get one FLAG peptide for free. Choice of FLAG peptide or FLAG-Cysteine peptide. Use Code FREEFLAG When Checkout.
Order DYKDDDDK peptide ($50 per mg) from here.
LifeTein's anti-DYKDDDDK monoclonal antibody was developed based on the most widely used hydrophilic octapeptide: DYKDDDDK. Order DYKDDDDK peptide ($50 per mg) from here. LifeTein's anti-DYKDDDDK renders higher affinity and higher binding capacity than Sigma's antibody. Please see the following table for technical details.
Comparison of anti-DYKDDDDK affinity gel with commercially available M2 affinity gel
|50 μg, lyophilized with PBS, pH7.4|
|Purchase matching Western Blot positive control protein|
|DYKDDDDK (FLAG*) synthetic peptide conjugated to KLH. Order DYKDDDDK peptide ($50 per mg) from here.|
|Recognizes the N-terminal, C-terminal or internal DYKDDDDK-tagged fusion proteins|
|Protein A affinity chromatography from mouse ascites fluid|
|Dot, ELISA, IS, IP, WB|
|WB (with ECL): 1-5ng of DYKDDDDK-tagged fusion proteins|
|WB (with ECL): 1:1000-3000 dilution (incubate for 1 hour at room temperature)|
IS: 1:500-2000 dilution
For best results with other assays (e.g.: Dot, ELISA, IP, etc), please determine optimal working dilution by titration test
|Store at -20°C for up to 2 years. Reconstitute the lyophilized antibody with deionized water (or equivalent) to a final concentration of 50ug/50ul. Aliquot to avoid repeated freeze-thaw cycles. Store aliquots at 4°C for 1-2 weeks |
Fig. 1:1000 (1µg/mL) Ab dilution probed against HEK293 cells transfected with DYKDDDDK-tagged protein vector; untransfected (1) and transfected (2)
Order DYKDDDDK peptide ($50 per mg) from here.
*FLAG is a registered trademark of Sigma-Aldrich Co. LLC
**ANTI-FLAG is a registered trademark of Sigma-Aldrich Co. LLC
1. Enzymology, Preexistent Asymmetry in the Human Cyclooxygenase-2 Sequence Homodimer, Liang Dong, Narayan P. Sharma, Brice J., Jurban and William L. Smith, J. Biol. Chem. published online August 16, 2013
Anti-FLAG monoclonal antibody was purchased from LifeTein to study the cyclooxygenase-2 (COX-2). It was found that the COX-2 can fold into a preexistent confromational heterodimer. COX-2 half-sites functionality results from COX-2 folding into stable conformational heterodimer.
... Hexane, isopropyl alcohol, and acetic acid were HPLC grade from Thermo Fisher Scientific, Inc.
Anti-PGHS-2 antibodies directed against the 18-amino acid insert unique to PGHS-2 were as
described (26). Anti-FLAG antibodies were from LifeTein, South Plainfield, NJ. ...
2. JM Eaton, GR Mullins, DN Brindley, TE Harris, Journal of Biological Chemistry, 2013, Phosphorylation of lipin 1 and charge on the phosphatidic acid head group control its phosphatidic acid phosphatase activity and membrane association, doi:10.1074/jbc.M112.441493jbc.M112.441493.
The FLAG sequence DYKDDDDK is frequently used for affinity purification. Synthetic FLAG peptide by LifeTein was used in the elution buffer to purify lipin 1 protein. It was found that the Lipin 1 preferentially binds di-anionic phosphatidic acid and this is eliminated by phosphorylation.
... The slurry was packed onto a screening column, washed with buffer A and lipin 1 was eluted by 5 successive additions of an equal volume of 0.5 mg/mL FLAG peptide (Lifetein) in 150 mM
NaCl, 20 mM Hepes, pH 7.2....
3. Brain tumor senescence might be mediated by downregulation of S-phase kinase-associated protein 2 via butylidenephthalide leading to decreased cell viability, 2014，1010-4283，J Tumor Biology.
Anti-FLAG monoclonal antibody was purchased from LifeTein to study the Brain tumor senescence. It was found that BP-triggered senescence in GBM cells is highly associated with its control on Skp2 regulation.
... DYKDDDDK Tag antibodies were purchased from LifeTein (South Plainfield, NJ, USA). ...
4. Methods for assaying protein-protein interactions, 2015, United States Patent Application
Anti-FLAG monoclonal antibody FG4R was purchased from LifeTein to study the protein-protein interactions by Western Blot.
... mouse anti-FLAG FG4R (LifeTein LT0420), ...
5. Arabidopsis BREVIPEDICELLUS Interacts with the SWI2/SNF2 Chromatin Remodeling ATPase BRAHMA to Regulate KNAT2 and KNAT6 Expression in Control of Inflorescence Architecture
Anti-FLAG monoclonal antibody was purchased from LifeTein to perform Co-IP Assays.
... Proteins were eluted with 40 μL of 2—loading buffer and analyzed by western blotting using ... anti-Flag antibodies (Life Tein) ...