Key Product Details

Species Reactivity

Epitope Tag

Applications

Validated:

ELISA, Flow Cytometry, Immunohistochemistry, Simple Western, Western Blot

Cited:

WB

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG2a Kappa Clone # 29E4.G7

Concentration

Please see the vial label for concentration. If unlisted please contact technical services.

View all DYKDDDDK Epitope Tag Antibodies »

Product Specifications

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.

Clonality

Monoclonal

Host

Mouse

Isotype

IgG2a Kappa

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.

Scientific Data Images for DYKDDDDK Epitope Tag Antibody (29E4.G7)

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.
View 1 more images

Applications for DYKDDDDK Epitope Tag Antibody (29E4.G7)

Application
Recommended Usage

ELISA

1:150000-1:250000

Immunohistochemistry

1:1000-1:5000

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.

Reviewed Applications

Read 3 reviews rated 5 using NBP1-97410 in the following applications:

Formulation, Preparation, and Storage

Purification

Protein A purified

Formulation

0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2

Preservative

0.01% Sodium Azide

Concentration

Please see the vial label for concentration. If unlisted please contact technical services.

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at -20C short term. Aliquot and store at -80C long term. Avoid freeze-thaw cycles.

Background: DYKDDDDK Epitope Tag

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.

Alternate Names

FLAG Tag

Additional DYKDDDDK Epitope Tag Products

Product Documents for DYKDDDDK Epitope Tag Antibody (29E4.G7)

Product Datasheets

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot number in the search box below.

Download SDS

Product Specific Notices for DYKDDDDK Epitope Tag Antibody (29E4.G7)

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.

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.

Privacy and Cookie Policy
Site Map
© Copyright 2024 Bio-Techne. All Rights Reserved.