Revision 1
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP, IF-IC

REACTIVITY:

All

SENSITIVITY:

Transfected Only

MW (kDa):

Source/Isotype:

Rabbit IgG

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:50
Immunofluorescence (Immunocytochemistry) 1:400 - 1:1600

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/mL BSA, 50% glycerol, and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

3X DYKDDDDK Tag (E7C5T) Rabbit mAb detects exogenously expressed proteins containing the 3X DYKDDDDK epitope tag. This antibody recognizes the 3X DYKDDDDK tag fused to either the amino or carboxy terminus of targeted proteins in transfected cells, but does not detect DYKDDDDK tag. 3X DYKDDDDK Tag (E7C5T) Rabbit mAb detects exogenously expressed 3X DYKDDDDK-tagged proteins in cells expressed under a CMV promoter. Expression under other promoters has not been evaluated.

Species Reactivity:

All Species Expected

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide containing the 3X DYKDDDDK epitope (DYKDHDGDYKDHDIDYKDDDDK).

Background

Epitope tags are useful for the labeling and detection of proteins using immunoblotting, immunoprecipitation, and immunostaining techniques. Because of their small size, they are unlikely to affect the tagged protein’s biochemical properties.

The DYKDDDDK peptide has been used extensively as a general epitope tag in expression vectors. This peptide can be expressed and detected with the protein of interest as an amino-terminal or carboxy-terminal fusion (1).

  1. Brizzard, B.L. et al. (1994) Biotechniques 16, 730-5.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation IF-IC: Immunofluorescence (Immunocytochemistry)

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Revision 1
#87537

3X DYKDDDDK Tag (E7C5T) Rabbit mAb

Western Blotting Image 1: 3X DYKDDDDK Tag (E7C5T) Rabbit mAb Expand Image
Western blot analysis of extracts from 293T cells, untransfected (-) or transfected with a construct expressing either an amino-terminal 3X DYKDDDDK-tagged GFP fusion protein (3X DYKDDDDK Tag-GFP; +) or carboxy-terminal 3X DYKDDDDK-tagged GFP fusion protein (GFP-3X DYKDDDDK Tag; +), using 3X DYKDDDDK Tag (E7C5T) Rabbit mAb (upper), GFP (D5.1) Rabbit mAb #2956 (middle), or β-Actin (D6A8) Rabbit mAb #8457 (lower).
Western Blotting Image 2: 3X DYKDDDDK Tag (E7C5T) Rabbit mAb Expand Image
Western blot analysis of extracts from control 293T cells (lane 1) or CRISPR/Cas9 3X DYKDDDDK-tagged WDR59 knockin (KI) 293T cells (lane 2) using 3X DYKDDDDK Tag (E7C5T) Rabbit mAb (upper) or WDR59 (D4Z7A) Rabbit mAb #53385 (lower).
Immunoprecipitation Image 1: 3X DYKDDDDK Tag (E7C5T) Rabbit mAb Expand Image
Immunoprecipitation of 3X DYKDDDDK Tag protein from CRISPR/Cas9 3X DYKDDDDK-tagged WDR59 knockin (KI) 293T cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is 3X DYKDDDDK Tag (E7C5T) Rabbit mAb. Western blot analysis was performed using WDR59 (D4Z7A) Rabbit mAb #53385. Anti-rabbit IgG, HRP-linked Antibody #7074 was used as a secondary antibody.
Immunoprecipitation Image 2: 3X DYKDDDDK Tag (E7C5T) Rabbit mAb Expand Image
Immunoprecipitation of 3X DYKDDDDK Tag protein from CRISPR/Cas9 3X DYKDDDDK-tagged WDR59 knockin (KI) 293T cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is 3X DYKDDDDK Tag (E7C5T) Rabbit mAb. Western blot analysis was performed using 3X DYKDDDDK Tag (E7C5T) Rabbit mAb. Anti-rabbit IgG, HRP-linked Antibody #7074 was used as a secondary antibody.
Immunofluorescence Image 1: 3X DYKDDDDK Tag (E7C5T) Rabbit mAb Expand Image
Confocal immunofluorescent analysis of CRISPR/Cas9 3X DYKDDDDK-tagged WDR59 knockin (KI) 293T cells (left, positive) and control 293T cells (right, negative) using 3X DYKDDDDK Tag (E7C5T) Rabbit mAb (green), DyLight 650 Phalloidin #12956 (red), and DAPI #4083 (blue).