DYKDDDDK Epitope Tag Antibody (29E4.G7) - BSA Free

Western Blot: DYKDDDDK Epitope Tag Antibody (29E4.G7) [NBP1-97410] - Use of DYKDDDDK Epitope Tag Antibody (29E4.G7) [NBP1-97410] in Western Blot. Flag expression is significantly increased in 293 cells transfected with a flag-fusion protein. Image from a verified customer review.

Western Blot: DYKDDDDK Epitope Tag Antibody (29E4.G7) [NBP1-97410] - Twenty-four clones were randomly selected and grown up from glycerol stocks by inoculating 0.5 mL 2X YT medium. Expression of recombinant proteins was induced by the addition of IPTG. Proteins were purified by nickel affinity chromatography and eluted in 40 uL. Samples were diluted 10-fold, transferred to nitrocellulose membrane and blotted using Mab DYKDDDDK Epitope Tag Antibody (29E4.G7) [NBP1-97410].

Western Blot: DYKDDDDK Epitope Tag Antibody (29E4.G7) [NBP1-97410] - Antibody detects both C-terminal linked and N-terminal linked FLAG tagged recombinant proteins.

ELISA: DYKDDDDK Epitope Tag Antibody (29E4.G7) [NBP1-97410] - Detection of DYKDDDDK conjugated proteins using the DyLight 405 conjugated version of DYKDDDDK Epitope Tag Antibody (29E4.G7).

• Reactivity: ET
• Applications: WB, Simple Western, ELISA, Flow, IHC
• Clone: 29E4.G7
• Clonality: Monoclonal
• Host: Mouse
• Conjugate: Unconjugated
• Format: BSA Free

DYKDDDDK Epitope Tag Antibody (29E4.G7) - BSA Free Summary

• Description: This product is an IgG fraction antibody purified from ascites by Protein A chromatography followed by extensive dialysis against the buffer stated above. The purified antibody is directed against the FLAG(TM) motif and is useful in determining its presence in various assays where the epitope tag is present at either the amino or carboxy terminus of recombinant proteins
  
  Store vial at -20C prior to opening. Aliquot contents and freeze at -20C or below for extended storage. Avoid cycles of freezing and thawing. Centrifuge product if not completely clear after standing at room temperature. This product is stable for several weeks at 4C as an undiluted liquid. Dilute only prior to immediate use.
• Immunogen: This DYKDDDDK Epitope Tag Antibody (29E4.G7) was produced in mice by repeated immunizations with a synthetic peptide corresponding to the FLAG(TM) epitope tag peptide DYKDDDDK Epitope Tag Epitope Tag (Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Lys) conjugated to KLH using maleimide. Residues of glycine and cysteine were added to the termini to facilitate coupling.
• Specificity: The purified antibody is directed against the FLAG(TM) motif and is useful in determining its presence in various assays where the epitope tag is present at either the amino or carboxy terminus of recombinant proteins. This monoclonal anti-FLAG(TM) tag antibody detects over-expressed proteins containing the FLAG(TM) epitope tag. In western blotting of bacterial extracts, the antibody does not cross-react with endogenous proteins.
• Isotype: IgG2a Kappa
• Clonality: Monoclonal
• Host: Mouse
• Purity: Protein A purified

Applications/Dilutions

• Dilutions:
  • ELISA 1:150000-1:250000
  • Flow Cytometry
  • Immunohistochemistry 1:1000-1:5000
  • Simple Western
  • Western Blot 1:2000-1:10000
• Application Notes: This product has been tested by ELISA and western blot and is optimally suited for monitoring the expression of FLAG(TM) tagged fusion proteins. As such, this antibody can be used to identify fusion proteins containing the FLAG(TM) epitope. The antibody recognizes the epitope tag fused to either the amino- or carboxy- termini of targeted proteins. The epitope tag peptide sequence was first derived from the 11-amino-acid leader peptide of the gene-10 product from bacteriophage T7. DYKDDDDK is the most commonly used hydrophilic octapeptide tag.
  
  Use in Western Blot and Simple Western reported in a Verified Customer Review. Dilution for use in Flow Cytrometry is user optimized.
  See Simple Western Antibody Database for Simple Western validation: separated by Size

Packaging, Storage & Formulations

• Storage: Store at -20C short term. Aliquot and store at -80C long term. Avoid freeze-thaw cycles.
• Buffer: 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2
• Preservative: 0.01% Sodium Azide
• Purity: Protein A purified

Notes

FLAG(R) and ANTI-FLAG(R) are registered trademarks of Sigma-Aldrich Co. LLC.

FLAG(R) and ANTI-FLAG(R) are registered trademarks of Sigma-Aldrich Co. LLC.

Alternate Names for DYKDDDDK Epitope Tag Antibody (29E4.G7) - BSA Free

• DDDDK epitope tag
• DDK
• DYKDDDDK Epitope Tag
• DYKDDDDK tag
• ECS epitope tag
• ECS tag
• Enterokinase Cleavage Site epitope tag
• Enterokinase Cleavage Site tag
• epitope tag flag
• FLAG (FLAG is a trademark of Sigma Aldrich)
• FLAG Tag
• Ty 1 Tag
• xxxDDDDK epitope tag

Background

Fusion proteins, in which a known peptide such as a His-Tag, GST-Tag, or DYKDDDDK (FLAG(R)) is cloned with the target of interest, were developed to simplify protein isolation. Since specific antibodies may not be available against the protein of interest, epitope tag antibodies are commonly used to detect protein expression.

The epitope tag, DYKDDDDK, is comprised of an eight amino acid sequence derived from the gene-10 product from bacteriophage T7 and contains an enterokinase-cleavage site (1). Its hydrophilic nature and short peptide sequence reduce possible interference with protein expression, post-translational modifications, protein interactions, and protein function (2). The DYKDDDDK (FLAG(R)) epitope tag can either be fused to the N-terminus, C-terminus, or placed at an internal position of the target protein, facilitating characterization of the protein of interest using immunodetection methods. Compared to the standard DYKDDDDK epitope tag antibody, the rat monoclonal DYKDDDDK Epitope Tag antibody (clone L5) is 2-8 times more sensitive (3).

References

1. Brizzard BL, Chubet RG, Vizard DL. (1994) Immunoaffinity purification of (FLAG(R)) epitope-tagged bacterial alkaline phosphatase using a novel monoclonal antibody and peptide elution. Biotechniques. 16(4):730-5. PMID: 8024796

2. Einhauer A, Jungbauer A. (2001) The (FLAG(R)) peptide, a versatile fusion tag for the purification of recombinant proteins. J Biochem Biophys Methods. 49(1-3):455-65. PMID: 11694294

3. Park SH, Cheong C, Idoyaga J, Kim JY, Choi JH, Do Y, Lee H, Jo JH, Oh YS, Im W, Steinman RM, Park CG. (2008) Generation and application of new rat monoclonal antibodies against synthetic DYKDDDDK Epitope Tag (FLAG(R)) and OLLAS tags for improved immunodetection. J Immunol Methods. 331(1-2):27-38. PMID: 18054954

FLAG(R) is a trademark of Sigma-Aldrich.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for DYKDDDDK Epitope Tag Antibody (NBP1-97410)(1)

Publication using NBP1-97410

• Zhang T, Cui S, Xiong X et al. PIH1D3-knockout rats exhibit full ciliopathy features and dysfunctional pre-assembly and loading of dynein arms in motile cilia Frontiers in cell and developmental biology 2023-10-12 [PMID: 37900281] (WB)

Reviews for DYKDDDDK Epitope Tag Antibody (NBP1-97410) (3)

Average Rating: 5

(Based on 3 reviews)

We have 3 reviews tested in 1 species: Human.

reviewed by: Verified Customer WB Human 01/08/2019

Summary

• Application: Western Blot
• Sample Tested: 293 cell extract
• Species: Human
• Lot: 33508

reviewed by: Verified Customer Simple Western Human 08/20/2015

Summary

• Application: Simple Western
• Sample Tested: HEK 293 cell lysate
• Species: Human

reviewed by: Grace zhou WB Human 07/30/2014

Summary

• Application: Western Blot
• Sample Tested: Hela whole cell lysate
• Species: Human

FAQs for DYKDDDDK Epitope Tag Antibody (NBP1-97410). (Showing 1 - 5 of 5 FAQ).

1. Are the FLAG Tag antibodies validated in Simple Western?
   • Yes, we offer 3 FLAG Tag antibodies that have been tested in Simple Western: NBP1-71705, NBP1-97410, and MAB8529.
2. What the theoretical molecular weight for FLAG Tag antibodies?
   • The TMW for DYKDDDDK epitope FLAG Tag is approximately 1.01 kDa.
3. Will FLAG Tag antibodies detect internal FLAG tags? My FLAG sequence is sandwiched between two other protein sequences.
   • If you are applying your sample(s) to SDS-PAGE, under reducing condition and follow by Western Blot, DYKDDDDK epitope tag antibodies should be able to detect the tag region located internally. However, it depends on the property of your chimeric protein and if your tag is in the hydrophobic or hydrophilic region. Although DYKDDDDK is very hydrophilic, it may not be exposed to the surface if embedded deeply inside the protein with a high content of hydrophobic amino acids with or without covalently-linked modifications such as lipid moieties.
4. Will FLAG Tag antibodies recognize FLAG at the C-terminal end?
   • Yes, FLAG Tag antibodies can recognize FLAG-tag at both the N- or C-terminal.
5. Do FLAG Tag antibodies comes in lyophilized form?
   • The following FLAG Tag antibodies come in lyophilized form: NBP1-97393, NBP1-97401, NBP1-97399, NBP1-97400, and NBP1-97402.