Recombinant Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-313611-anti-ddddk-tag-antibody-epr20018-251-equivalent-to-flag-antibodies-from-sigma-immunohistochemistry.jpg "Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)")
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR20018-251] to DDDDK tag (Binds to FLAG® tag sequence)
- Suitable for: WB, ICC/IF, Flow Cyt, IHC-P, IP
- Reacts with: Species independent
Related conjugates and formulations
Overview
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Product name
Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251]
See all DDDDK tag (Binds to FLAG® tag sequence) primary antibodies -
Description
Rabbit monoclonal [EPR20018-251] to DDDDK tag (Binds to FLAG® tag sequence) -
Host species
Rabbit -
Tested applications
Suitable for: WB, ICC/IF, Flow Cyt, IHC-P, IPmore details -
Species reactivity
Reacts with: Species independent -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: HEK-293 transfected with PFKFB3 and HEK-293 transfected with PD-L1 cell lysate. IHC-P: HEK-293T transfected with DDDDK-tagged human PD-L1. ICC/IF: HEK-293T cells. Flow: HEK-293T transfected with DDDDK-tagged human PD-L1. IP: HEK-293T transfected with DDDDK-tagged human PFKFB3.
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General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
FLAG® is a registered trade mark of Sigma Aldrich Biotechnology LP. It is used here for informational purposes only.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading... -
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR20018-251 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab205606 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Abreviews | Notes |
|---|---|---|
| WB | (1) |
1/500 - 1/5000.
This antibody can be used at 1.2 μg/ml. ab205606 can detect extra bands in non-transfected cell lines at low dilution. |
| ICC/IF |
1/100.
|
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| Flow Cyt |
1/700.
|
|
| IHC-P |
1/750. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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| IP |
1/30.
|
| Notes |
|---|
|
WB
1/500 - 1/5000. This antibody can be used at 1.2 μg/ml. ab205606 can detect extra bands in non-transfected cell lines at low dilution. |
|
ICC/IF
1/100. |
|
Flow Cyt
1/700. |
|
IHC-P
1/750. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
|
IP
1/30. |
Target
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Relevance
This is a useful tool for the localisation and characterisation of DDDDK tagged proteins (Binds to FLAG® tag sequence). -
Alternative names
- DDDDK epitope tag antibody
- DDDK antibody
- ddk antibody
see all
Images
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)
Immunhistochemical analysis of agarose-embedded HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) transfected with DDDDK-tagged human PD-L1 expression vector labeling DDDDK tag with ab205606 at 1/750 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Counterstained with hematoxylin.
Positive staining on HEK-293T cells transfected with DDDDK-tagged human PD-L1 expression vector (Panel A) is observed. No signal was detected on HEK-293T transfected with an empty vector (vector control), containing a C-terminal DDDDK tag (Panel B).
Heat mediated antigen retrieval was performed using EDTA buffer pH 9 before commencing with IHC staining protocol.
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![Immunocytochemistry/ Immunofluorescence - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-313615-anti-ddddk-tag-antibody-epr20018-251-equivalent-to-flag-antibodies-from-sigma-immunofluorescence.jpg)
Immunocytochemistry/ Immunofluorescence - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) cells labeling DDDDK tag with ab205606 at 1/100 dilution, followed by ab150077 AlexaFluor®488 Goat anti-Rabbit secondary at 1/1000 dilution (green).
Confocal image showing positive staining for FLAG® on HEK-293T cells transfected with DDDDK-tagged PFKFB3 expression vector. Mouse monoclonal ANTI-FLAG® M2 antibody was used as a counterstain at 1/500 dilution, and AlexaFluor®647 Goat anti-mouse secondary (ab150115) was used as the secondary antibody only control at 1/500 dilution. The nucleus is counterstained with DAPI.
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![Western blot - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-313608-anti-ddddk-tag-antibody-epr20018-251-equivalent-to-flag-antibodies-from-sigma-western-blot.jpg)
Western blot - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)All lanes : Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606) at 1/1000 dilution
Lane 1 : HEK-293 (human epithelial cell line from embryonic kidney) transfected with an empty vector (vector control), containing an N-terminal DDDDK tag, whole cell lysate
Lane 2 : HEK-293 transfected with PFKFB3 (WT) expression vector containing an N-terminal DDDDK tag, whole cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Observed band size: 60 kDa why is the actual band size different from the predicted?Blocking/Diluting buffer: 5% NFDM/TBST
Exposure time: 0.5 seconds
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![Western blot - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-313609-anti-ddddk-tag-antibody-epr20018-251-equivalent-to-flag-antibodies-from-sigma-western-blot.jpg)
Western blot - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)All lanes : Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606) at 1/5000 dilution
Lane 1 : HEK-293 (human epithelial cell line from embryonic kidney) transfected with an empty vector (vector control), containing a C-terminal DDDDK tag, whole cell lysate
Lane 2 : HEK-293 transfected with PD-L1 (WT) expression vector containing a C- terminal DDDDK tag, whole cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilutionBlocking/Diluting buffer: 5% NFDM/TBST
Exposure time: 1 second
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![Immunoprecipitation - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-313617-anti-ddddk-tag-antibody-epr20018-251-equivalent-to-flag-antibodies-from-sigma-immunoprecipitation.jpg)
Immunoprecipitation - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)DDDDK tag was immunoprecipitated from 0.35 mg HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) transfected with DDDDK-tagged human PFKFB3 expression vector whole cell lysate with ab205606 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab205606 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.
Lane 1: HEK-293T transfected with DDDDK-tagged human PFKFB3 expression vector whole cell lysate (input).
Lane 2: ab205606 IP in HEK-293T transfected with DDDDK-tagged human PFKFB3 expression vector whole cell lysate.Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab205606 in HEK-293T transfected with DDDDK-tagged human PFKFB3 expression vector whole cell lysate.
Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 3 seconds. -
![Western blot - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-380983-anti-ddddk-tag-antibody-epr20018251-western-blot-hek293-nih3t3-c6-human-mouse-rat.jpg)
Western blot - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)All lanes : Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606) at 1.2 µg/ml (1:500 dilution)
Lane 1 : HEK-293 (human epithelial cell line from embryonic kidney) whole cell lysate
Lane 2 : NIH/3T3 (mouse embryo fibroblast cell line) whole cell lysate
Lane 3 : C6 (rat glial tumor cell line) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)
Developed using the ECL technique.
Observed band size: 25,30,45,80 kDa why is the actual band size different from the predicted?
Exposure time: 20 secondsBlocking/Diluting buffer and concentration: 5% NFDM/TBST.
ab205606 can detect extra bands in non-transfected cell lines at low dilution.
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![Flow Cytometry - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-313616-anti-ddddk-tag-antibody-epr20018-251-equivalent-to-flag-antibodies-from-sigma-flow-cytometry.jpg)
Flow Cytometry - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)Flow cytometric analysis of of 4% paraformaldehyde-fixed, 90% methanol permeabilized HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) transfected with DDDDK-tagged human PD-L1 expression vector labeling DDDDK tag with ab205606 at 1/700 dilution (Red) compared with the Rabbit monoclonal IgG isotype control (ab172730) (Black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.
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![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-313614-anti-ddddk-tag-antibody-epr20018-251-equivalent-to-flag-antibodies-from-sigma-immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)Negative control: No staining on rat spleen.
Immunohistochemical analysis of paraffin-embedded rat spleen tissue stained for DDDDK tag using ab205606 at 1/750 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Counterstained with hematoxylin.
Heat mediated antigen retrieval was performed using EDTA buffer pH 9 before commencing with IHC staining protocol.
-
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-313613-anti-ddddk-tag-antibody-epr20018-251-equivalent-to-flag-antibodies-from-sigma-immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)Negative control: No staining on mouse spleen.
Immunohistochemical analysis of paraffin-embedded mouse spleen tissue stained for DDDDK tag using ab205606 at 1/750 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Counterstained with hematoxylin.
Heat mediated antigen retrieval was performed using EDTA buffer pH 9 before commencing with IHC staining protocol.
-
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-313612-anti-ddddk-tag-antibody-epr20018-251-equivalent-to-flag-antibodies-from-sigma-immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)Negative control: No staining on human hepatocellular cancer.
Immunohistochemical analysis of paraffin-embedded human hepatocellular cancer tissue stained for DDDDK tag using ab205606 at 1/750 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Counterstained with hematoxylin.
Heat mediated antigen retrieval was performed using EDTA buffer pH 9 before commencing with IHC staining protocol.
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![Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)](https://www.abcam.com/ps/products/205/ab205606/Images/ab205606-11-benefits-of-recombinant-antibodies.png)
Anti-DDDDK tag (Binds to FLAG® tag sequence) antibody [EPR20018-251] (ab205606)
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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SDS download
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Datasheet download
Certificate of Compliance
References (29)
ab205606 has been referenced in 29 publications.
- Yan G et al. Deubiquitylation and stabilization of ARMC5 by ubiquitin-specific processing protease 7 (USP7) are critical for RCC proliferation. J Cell Mol Med N/A:N/A (2021). PubMed: 33544460
- Zheng F et al. The HIF-1a antisense long non-coding RNA drives a positive feedback loop of HIF-1a mediated transactivation and glycolysis. Nat Commun 12:1341 (2021). PubMed: 33637716
- Lu X et al. The miRNA-15b/USP7/KDM6B axis engages in the initiation of osteoporosis by modulating osteoblast differentiation and autophagy. J Cell Mol Med 25:2069-2081 (2021). PubMed: 33434305
- Guo ZF & Kong FL Akt regulates RSK2 to alter phosphorylation level of H2A.X in breast cancer. Oncol Lett 21:187 (2021). PubMed: 33574926
- Lei G et al. TRIM29 Reverses Oxaliplatin Resistance of P53 Mutant Colon Cancer Cell. Can J Gastroenterol Hepatol 2021:8870907 (2021). PubMed: 33824865