DYKDDDDK tag Polyclonal antibody (Binds to FLAG® tag epitope)

DYKDDDDK tag Polyclonal Antibody for WB, ELISA

Host / Isotype

Rabbit / IgG

Reactivity

recombinant protein and More (8)

Applications

WB, RIP, IP, IHC, IF, FC, CoIP, ChIP, ELISA

Conjugate

Unconjugated

Cat no : 20543-1-AP

Synonyms

DDDDK, DDDDK tag, DYKDDDDK, DYKDDDDK tag, Flag


Filter:
  • Western Blot (WB) analysis of various lysates using DYKDDDDK tag Polyclonal antibody (Binds to FLAG® t (20543-1-AP)

    WB analysis using 20543-1-AP

    HEK-293T cells and transfected HEK-293T lysates were subjected to SDS PAGE followed by western blot with 20543-1-AP (DYKDDDDK tag antibody) at dilution of 1:50000 incubated at room temperature for 1.5 hours.

"DYKDDDDK tag Antibodies" Comparison

View side-by-side comparison of DYKDDDDK tag antibodies from other vendors to find the one that best suits your research needs.

Tested Applications

Positive WB detected inTransfected HEK-293T cells

Recommended dilution

ApplicationDilution
Western Blot (WB)WB : 1:20000-1:100000
It is recommended that this reagent should be titrated in each testing system to obtain optimal results.
Sample-dependent, Check data in validation data gallery.

Product Information

20543-1-AP targets DYKDDDDK tag in WB, RIP, IP, IHC, IF, FC, CoIP, ChIP, ELISA applications and shows reactivity with recombinant protein samples.

Tested Reactivity recombinant protein
Cited Reactivityhuman, mouse, rat, chicken, yeast, monkey, pig, duck
Host / Isotype Rabbit / IgG
Class Polyclonal
Type Antibody
Immunogen DYKDDDDK tag fusion protein Ag2329
Full Name DYKDDDDK tag
Gene Symbol
Gene ID (NCBI)
RRIDAB_11232216
Conjugate Unconjugated
Form Liquid
Purification MethodAntigen affinity purification
Storage Buffer PBS with 0.02% sodium azide and 50% glycerol pH 7.3.
Storage ConditionsStore at -20°C. Stable for one year after shipment. Aliquoting is unnecessary for -20oC storage. 20ul sizes contain 0.1% BSA.

Background Information

Protein tags are protein or peptide sequences located either on the C- or N- terminal of the target protein, which facilitates one or several of the following characteristics: solubility, detection, purification, localization and expression. The DYKDDDDK(FLAG) peptide has been used extensively as a general tag in expression vectors. This peptide can be expressed and detected with the protein of interest as an amino-terminal or carboxy-terminal fusion. N-terminal DDDDK vectors provide an Ek cleavage site for removal of the fusion tag. The DDDDK peptide is likely to be located on the surface of a fusion protein because of its hydrophilic nature. As a result, the DDDDK peptide is more likely to be accessible to antibodies. A DDDDK-tag can be used in many different assays that require recognition by an antibody, such as western blotting, immunocytochemistry, immunoprecipitation, flow cytometry, protein purification, and in the study of protein-protein interactions, cell ultrastructure, and protein localization and so on. This antibody is a rabbit polyclonal antibody raised against 3xFlag (3xDYKDDDDKT) sequence and recognizes the (1x) and (3x)DYKDDDDK peptide and detects DDDDK-tagged proteins. Anti-FLAG is a registered trademark of Sigma-Aldrich Biotechnology.

Protocols

Product Specific Protocols
WB protocol for DYKDDDDK tag antibody 20543-1-APDownload protocol
Standard Protocols
Click here to view our Standard Protocols

Publications

SpeciesApplicationTitle
IP

Science

Ubiquitination of G3BP1 mediates stress granule disassembly in a context-specific manner.

Authors - Youngdae Gwon
IP

Cell

Ca2+ sensor-mediated ROS scavenging suppresses rice immunity and is exploited by a fungal effector.

Authors - Mingjun Gao
WB,IP

Nat Metab

Lactylome analysis suggests lactylation-dependent mechanisms of metabolic adaptation in hepatocellular carcinoma

Authors - Zijian Yang
WB

Cell Res

Cholesterylation of Smoothened is a calcium-accelerated autoreaction involving an intramolecular ester intermediate.

Authors - Ao Hu

Cell Res

Oncogenic AURKA-enhanced N6-methyladenosine modification increases DROSHA mRNA stability to transactivate STC1 in breast cancer stem-like cells.

Authors - Fei Peng
WB,CoIP

Crit Care

Recombinant ACE2 protein protects against acute lung injury induced by SARS-CoV-2 spike RBD protein.

Authors - Lingbing Zhang

Reviews

The reviews below have been submitted by verified Proteintech customers who received an incentive forproviding their feedback.


FH

Jon (Verified Customer) (02-28-2023)

The antibody works fine in WB in the range fo 1:1000-1:5000 dilution in 5% milk TBST. In the image the samples are HEK293 lysates on a tricine SDS PAGE with primary antiFLAG antibody 1:5000 overnight at 4 ºC. It is true that the antibody could show considerable nonspecific background depending on the concentrations and cells used. In my experience, works perfectly fine and is quite sensitive in HEK293 cells, although in my EndoC-ßH1 cells signal is weaker and backgorund is very notable. However, I assume it is due to the EndoC cells, which are a pain for many applications. I also used this antibody for some IF experiments. In the EndoC line, no matter the antibody concentration, I always get non specific signal. I'm worried the antibody binds to something in particular in these cells. In HEK293 cells however, I managed to obtain good results for overexpression of a FLAG tagged peptide, same as the one on Western Blot

  • Applications: Western Blot, Immunofluorescence
  • Primary Antibody Dilution: For WB 1:2000 and for IF 1:1000
  • Cell Tissue Type: HEK293
DYKDDDDK tag Antibody Western Blot,Immunofluorescence validation (For WB 1:2000 and for IF 1:1000 dilution) in HEK293 (Cat no:20543-1-AP)
FH

Brogan (Verified Customer) (11-09-2022)

Also used in a pul down assay for FLAG tag plasmids

  • Applications: Western Blot, Immunoprecipitation, Protein purification
FH

Chun (Verified Customer) (06-19-2022)

This antibody worked very well in both ECL and Li-Cor procedures.

  • Applications: Western Blot
  • Primary Antibody Dilution: 1:1,000
  • Cell Tissue Type: HeLa cells and HEK293 cells transfected with plasmids encoding Flag-tagged proteins
FH

WEI (Verified Customer) (05-11-2022)

Good for western blot of knock-in mice but not good for immunostaining.

  • Applications: Western Blot
  • Cell Tissue Type: Mouse Heart
FH

Barbara (Verified Customer) (03-28-2022)

reliable antibody for detection of FLAG tag by western blot

  • Applications: Western Blot
  • Primary Antibody Dilution: 1:1000
  • Cell Tissue Type: Neuro-2A lysate
FH

ISABELLE CRISTINE (Verified Customer) (01-14-2022)

This antibody is extremely concentrated and works very well for detecting FLAG-tagged proteins in U2OS and HEK293 cells.

  • Applications: Western Blot
  • Primary Antibody Dilution: 1:4000
  • Cell Tissue Type: HEK293 and U2OS cells
DYKDDDDK tag Antibody Western Blot validation (1:4000 dilution) in HEK293 and U2OS cells (Cat no:20543-1-AP)
FH

X (Verified Customer) (01-05-2022)

It is Ok to use it in WB but with some non-specific bands.

  • Applications: Western Blot
  • Primary Antibody Dilution: 1:2000
  • Cell Tissue Type: primary cortical neuron infected with FLAG-fusion protein expressing lentivirus
DYKDDDDK tag Antibody Western Blot validation (1:2000 dilution) in primary cortical neuron infected with FLAG-fusion protein expressing lentivirus (Cat no:20543-1-AP)
FH

Christine (Verified Customer) (10-13-2021)

Antibody used on HEK293T transfected with a FLAG tagged protein and western blots compared with 3 other FLAG antibodies. This one gives the strongest signal for the tagged protein (which is a about 34 kDa) so it might be very useful for poorly expressed proteins. However it also has the most background bands, with 2 main additional background bands (but with a lower intensity than my tagged protein of interest): one at around 50 kDa (just above 47 kDa marker) and another one between 118 and 85 kDa markers. So this antibody is worth a try depending on the size of your protein of interest. It gives the strongest signal of the 4 antibodies tested.

  • Applications: Western Blot
  • Primary Antibody Dilution: 1:1000
  • Cell Tissue Type: Transfected HEK293T cell line
FH

Boyan (Verified Customer) (02-05-2021)

Very strong signal, but with some weak non specific bands.

  • Applications: Western Blot
FH

Lahiru Chamara (Verified Customer) (11-06-2020)

HEK293 cells were transfected with gene of interested. Primary antibody incubation was performed at 4C for an overnight and Alexa Fluor 488 secondary antibody incubation was performed at room temperature for an hour. Cells were stained with Hoechst for DNA/ nuclei . Images were taken from Zeiss LSM780 microscope.

  • Applications: Western Blot, Immunohistochemistry
  • Primary Antibody Dilution: 1:500
  • Cell Tissue Type: Transfected HEK293
DYKDDDDK tag Antibody Western Blot,Immunohistochemistry validation (1:500 dilution) in Transfected HEK293 (Cat no:20543-1-AP)
FH

Tongbin (Verified Customer) (08-25-2020)

For western blots, this FLAG antibody is comparable to Sigma's M2 FLAG antibody, which has been the gold standard for FLAG.

  • Applications: Western Blot
  • Primary Antibody Dilution: 1:1000
  • Cell Tissue Type: HEK293T
FH

Dipen (Verified Customer) (11-11-2019)

Great antibody for Western blotting. I used it at 1:1000 and got a super strong signal. Will use at a lower concentration next time.

  • Applications: Western Blot,
  • Primary Antibody Dilution: 1:1000
  • Cell Tissue Type: HEK293T cells
DYKDDDDK tag Antibody Western Blot, validation (1:1000 dilution) in HEK293T cells (Cat no:20543-1-AP)
FH

Sara (Verified Customer) (10-24-2019)

Works really well for both IP of overexpressed FLAG-tagged proteins and WB!

  • Applications: Western Blot, Immunoprecipitation,
  • Primary Antibody Dilution: 1:2000
  • Cell Tissue Type: 293T overexpressing FLAG-tagged construct
DYKDDDDK tag Antibody Western Blot,Immunoprecipitation, validation (1:2000 dilution) in 293T overexpressing FLAG-tagged construct (Cat no:20543-1-AP)
FH

Nikhil (Verified Customer) (10-16-2019)

Flag tagged p53 expressed in H1299 cells.

  • Applications: Western Blot,
  • Primary Antibody Dilution: 1:1000
  • Cell Tissue Type: H1299
DYKDDDDK tag Antibody Western Blot, validation (1:1000 dilution) in H1299 (Cat no:20543-1-AP)
FH

Shivani (Verified Customer) (09-26-2019)

Works better than another brand I had tested.

  • Applications: Western Blot,
  • Primary Antibody Dilution: 1:1000
FH

Corey (Verified Customer) (08-23-2019)

This antibody has worked well for western blotting as well as immunoprecipitation. The lots have been consistent so far, we will definitely be ordering more.

  • Applications: Western Blot, Immunoprecipitation,
  • Primary Antibody Dilution: 1:1000
  • Cell Tissue Type: H1299
FH

Kishor (Verified Customer) (06-23-2019)

Works very good for WB at 1:1000 and IP at 1:200.

  • Applications: Western Blot, Immunoprecipitation,
  • Primary Antibody Dilution: 1:1000
  • Cell Tissue Type: Human cell lines
FH

David (Verified Customer) (06-19-2019)

This is working very well for me. I ran out of another brand and got this one because I saw you had it and it was a better value. After testing it in the lab, it works just as well, if not better than the previous one. A very strong signal in my experiments.

  • Applications: Western Blot,
  • Primary Antibody Dilution: 1/2000
FH

Lana (Verified Customer) (04-20-2019)

SDS-PAGE:10 ug/ul RIPA lysate of whole cells4-12% Bis-tris gradient gelTransfer:Immobilon-FL transfer membranes (Millipore) O/N at 30V, 4CBlocking:SEA Block Blocking Buffer 1hPrimary Ab:DDDDK pRb Cat# 20543-1-APO/N incubation at 4CSecondary Ab:IRDye 800CW Goat anti-Rabbit1 h incubation at room temperatureLines on WB:1. BioRad Precision Plus Protein standards2. Whole cell lysate of HEK293t cells (negative transfection)3. Whole cell lysate of HEK293t cells transfected with FLAG-tag construct 27 kDa3. Whole cell lysate of HEK293t cells transfected with FLAG-tag construct 40 kDa

  • Applications: Western Blot,
  • Primary Antibody Dilution: 1:1000
  • Cell Tissue Type: HEK293t
DYKDDDDK tag Antibody Western Blot, validation (1:1000 dilution) in HEK293t (Cat no:20543-1-AP)
FH

Luce (Verified Customer) (11-12-2018)

  • Applications: Western Blot, Immunoprecipitation,
  • Cell Tissue Type: 10T1/2 cells