Name: Anti-TRAF6 antibody [EP591Y]
Brand: RabMAb
SKU: ab33915
Rating: 4 (9 reviews)

Product name

Anti-TRAF6 antibody [EP591Y]
See all TRAF6 primary antibodies
Description

Rabbit monoclonal [EP591Y] to TRAF6
Host species

Rabbit
Tested applications

Suitable for: WB, Flow Cyt, IHC-Pmore details
Unsuitable for: IP
Species reactivity

Reacts with: Mouse, Rat, Human
Predicted to work with: Zebrafish
Immunogen

Synthetic peptide within Human TRAF6 aa 50-150 (N terminal). The exact sequence is proprietary.
(Peptide available as ab183540)
Positive control
- Jurkat and HAP1 whole cell lysate. Human colon adenocarcinoma. This antibody is unsuitable for detecting TRAF6 in tissue lysates.
General notes

Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab^® patents.
This product is a recombinant rabbit monoclonal antibody.

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
Storage buffer

pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 49% PBS, 50% Glycerol, 0.05% BSA
Concentration information loading...
Purity

IgG fraction
Clonality

Monoclonal
Clone number

EP591Y
Isotype

IgG
Research areas

Alternative Versions
Compatible Secondaries
Immunizing Peptide (Blocking)
- TRAF6 peptide (ab183540)
Isotype control
- Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
Positive Controls
- Jurkat whole cell lysate (ab7899)

Our Abpromise guarantee covers the use of ab33915 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		1/2000 - 1/10000. Predicted molecular weight: 58 kDa.Can be blocked with TRAF6 peptide (ab183540).
Flow Cyt		1/50. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
IHC-P		Use at an assay dependent concentration.

Application notes

Is unsuitable for IP.

Function

E3 ubiquitin ligase that, together with UBE2N and UBE2V1, mediates the synthesis of 'Lys-63'-linked-polyubiquitin chains conjugated to proteins, such as IKBKG, AKT1 and AKT2. Also mediates ubiquitination of free/unanchored polyubiquitin chain that leads to MAP3K7 activation. Leads to the activation of NF-kappa-B and JUN. May be essential for the formation of functional osteoclasts. Seems to also play a role in dendritic cells (DCs) maturation and/or activation. Represses c-Myb-mediated transactivation, in B lymphocytes. Adapter protein that seems to play a role in signal transduction initiated via TNF receptor, IL-1 receptor and IL-17 receptor.
Tissue specificity

Expressed in heart, brain, placenta, lung, liver, skeletal muscle, kidney and pancreas.
Pathway

Protein modification; protein ubiquitination.
Sequence similarities

Belongs to the TNF receptor-associated factor family. A subfamily.
Contains 1 MATH domain.
Contains 1 RING-type zinc finger.
Contains 2 TRAF-type zinc fingers.
Domain

The coiled coil domain mediates homo- and hetero-oligomerization.
The MATH/TRAF domain binds to receptor cytoplasmic domains.
Post-translational
modifications

Sumoylated on Lys-124, Lys-142 and Lys-453 by SUMO1.
Polyubiquitinated on Lys-124; after cell stimulation with IL-1-beta or TGF-beta. This ligand-induced cell stimulation leads to dimerization/oligomerization of TRAF6 molecules, followed by auto-ubiquitination which involves UBE2N and UBE2V1 and leads to TRAF6 activation. This 'Lys-63' site-specific poly-ubiquitination appears to be associated with the activation of signaling molecules. Endogenous autoubiquitination occurs only for the cytoplasmic form.
Cellular localization

Cytoplasm. Cytoplasm > cell cortex. Nucleus. Found in the nuclei of some agressive B-cell lymphoma cell lines as well as in the nuclei of both resting and activated T-and B-lymphocytes. Found in punctate nuclear body protein complexes. Ubiquitination may occur in the cytoplasm and sumoylation in the nucleus.
Target information above from: UniProt accession Q9Y4K3 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 7189 Human
- Entrez Gene: 22034 Mouse
- Entrez Gene: 311245 Rat
- Entrez Gene: 554561 Zebrafish
- Omim: 602355 Human
- SwissProt: Q9Y4K3 Human
- SwissProt: P70196 Mouse
- SwissProt: B5DF45 Rat
- SwissProt: Q6IWL4 Zebrafish
- Unigene: 444172 Human
- Unigene: 591983 Human
- Unigene: 292729 Mouse
- Unigene: 220435 Rat
- Unigene: 74618 Zebrafish
see all
Alternative names
- E3 ubiquitin-protein ligase TRAF6 antibody
- Interleukin 1 signal transducer antibody
- Interleukin-1 signal transducer antibody
- MGC 3310 antibody
- MGC:3310 antibody
- MGC3310 antibody
- OTTHUMP00000232772 antibody
- OTTHUMP00000232773 antibody
- RING finger protein 85 antibody
- RNF 85 antibody
- RNF85 antibody
- TNF receptor associated factor 6 antibody
- TNF receptor-associated factor 6 antibody
- TNF receptor-associated factor 6, E3 ubiquitin protein ligase antibody
- TRAF 6 antibody
- Traf6 antibody
- TRAF6_HUMAN antibody
see all

Western blot - Anti-TRAF6 antibody [EP591Y] (ab33915)

Lane 1: Wild type HAP1 whole cell lysate (20 µg)
Lane 2: TRAF6 knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: HEK293 whole cell lysate (20 µg)

Lanes 1 - 4: Merged signal (red and green). Green - ab33915 observed at 65 kDa. Red - loading control, ab8245, observed at 37 kDa.

Ab33915 was shown to specifically react with TRAF6 in wild-type cells as signal was lost in TRAF6 knockout HAP1 cells. Wild-type and TRAF6 knockout samples were subjected to SDS-PAGE. Ab33915 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 2000 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging
Flow Cytometry - Anti-TRAF6 antibody [EP591Y] (ab33915)

Overlay histogram showing HAP1 wildtype (green line) and HAP1-TRAF6 knockout cells (red line) stained with ab33915. The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab33915, 0.1µg/ml ) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) preadsorbed (ab150081) at 1/2000 dilution for 30 min at 22°C.

A rabbit IgG isotype control antibody (ab172730) was used at the same concentration and conditions as the primary antibody (HAP1 wildtype - black line, HAP1-TRAF6 knockout - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).

Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TRAF6 antibody [EP591Y] (ab33915)

Immunohistochemical analysis of paraffin-embedded human colon adenocarcinoma using anti-TRAF6 (ab33915)
Flow Cytometry - Anti-TRAF6 antibody [EP591Y] (ab33915)Image courtesy of an anonymous Abreview.

ab33915 staining TRAF6 in Human platelet cells by Flow cytometry.
Cells were fixed in paraformaldehyde and permeabilized using 0.1% Triton-X-100 in 2% BSA for 15 minutes. Primary antibody used at a 1/200 dilution and incubated for 16 hours at 4°C. The secondary antibody used was an Alexa Fluor®488 conjugated chicken anti-rabbit IgG (H+L) at a 1/500 dilution.
P : Permeabilized
US : Unstained (Red Peak)
IGG RB : IgG Rabbit (Blue Peak)
TRAF6 Ab (Green Peak)
See Abreview
Western blot - Anti-TRAF6 antibody [EP591Y] (ab33915)

Anti-TRAF6 antibody [EP591Y] (ab33915) at 1/10000 dilution + Jurkat cell lysate

Predicted band size: 58 kDa
Flow Cytometry - Anti-TRAF6 antibody [EP591Y] (ab33915)

Flow Cytometry analysis of Jurkat (human acute T cell leukemia) cells labeling TRAF6 with purified ab33915 at 1/240 dilution(10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488)(1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) was used as the unlabeled control.
Western blot - Anti-TRAF6 antibody [EP591Y] (ab33915)

All lanes : Anti-TRAF6 antibody [EP591Y] (ab33915) at 1/1000 dilution

Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
Lane 2 : Human heart lysates
Lane 3 : Human skeletal muscle lysates
Lane 4 : Mouse skeletal muscle lysates
Lane 5 : Rat skeletal muscle lysates

Lysates/proteins at 15 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)

Predicted band size: 58 kDa

This antibody is unsuitable for detecting tissue lysates.
Western blot - Anti-TRAF6 antibody [EP591Y] (ab33915)

All lanes : Anti-TRAF6 antibody [EP591Y] (ab33915) at 1/1000 dilution

Lane 1 : Raw264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates
Lane 2 : PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysates

Lysates/proteins at 15 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)

Predicted band size: 58 kDa

This product has been referenced in:

Jang JH et al. Molecular cloning and functional characterization of TRAF6 and TAK1 in rainbow trout, Oncorhynchus mykiss. Fish Shellfish Immunol 84:927-936 (2019). Read more (PubMed: 30391533) »
DE Moura CFG et al. Evaluation of the Chemopreventive Activity of Grape Skin Extract Using Medium-term Oral Carcinogenesis Assay Induced by 4-Nitroquinoline 1-Oxide. Anticancer Res 39:177-182 (2019). Read more (PubMed: 30591456) »

See all 31 Publications for this product

Publishing research using ab33915? Please let us know so that we can cite the reference in this datasheet.

Customer reviews and Q&As

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1-10 of 16 Abreviews or Q&A

Application

Western blot

Loading amount

25 µg

Gel Running Conditions

Reduced Denaturing

Sample

Mouse Cell lysate - whole cell (liver)

Specification

liver

Blocking step

Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C

Abcam user community

Verified customer

Submitted Nov 17 2014

Application

Western blot

Loading amount

7.5 µg

Gel Running Conditions

Non-reduced Denaturing (7.5%)

Sample

Zebrafish Tissue lysate - whole (embryos 5-7 days post fertilization)

Specification

embryos 5-7 days post fertilization

Blocking step

BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 23°C

Rub�n Valued Customer

Verified customer

Submitted Oct 22 2013

Question

Is this antibody truly a monoclonal? (I have never heard of a rabbitmonoclonal.)
https://www.abcam.com/TRAF6-antibody-EP591Y-ab33915.html

Abcam community

Verified customer

Asked on Mar 26 2012

Answer

Thank you for contacting Abcam and for your interest in ab33915.

I can confirm for you that this is indeed a rabbit monoclonal antibody. Rabbit antibodies are generally high affinity and are an ideal species to choose for making antibodies against mouse and rat proteins as many antibodies made in mice or rats will not react against mouse or rat proteins. From our experiences the rabbit immune system appears to be capable of generating affinities against small epitopes which are not recognized by the mouse.

I hope this information is helpful. Please do not hesitate to contact us if you have any additional questions or concerns.

Abcam Scientific Support

Answered on Mar 26 2012

Question

Can you recommend EasyLink conjugation kits for bothe IRAK1 and TRAF6 kits, please.

Abcam community

Verified customer

Asked on Feb 23 2012

Answer

Thank you for your reply.

Our IRAK antibody ab238 will be compatible with any of our EasyLink conjugation kits. The full list of available sizes and fluorophores can be found at https://www.abcam.com/EasyLink, but for example you could use the FITC conjugation kit ab102885.

The TRAF6 antibody ab33915 is a rabbit monoclonal. Because the binding affinity of rabbit IgG is much, much higher than that of mouse IgG, a rabbit monoclonal antibody will be suitable for use at a far lower concentration. For that reason, ab33915 is sold at a very low concentration. The EasyLink kits can only be used with greater than 10ug of antibody per conjugation, in a volume of 4-10ul. That being said, even with the low concentration, we will guarantee this antibody to work at a dilution of 1:50 for Flow Cytometry.

I hope this helps, please let me know if you need any other information.

Abcam Scientific Support

Answered on Feb 23 2012

Question

I would like to know whether there are ELISA kits available for measuring TRAF6, IRAK1, STAT1 and IRF5 in serum?

Abcam community

Verified customer

Asked on Feb 14 2012

Answer

Thank you for contacting us.

We do not currently have any ELISA kits to TRAF6, IRAK1, or IRF5. We currently have three ELISA kits for STAT1 phosphorylations, but none to total STAT1:

ab119649: PhosphoTracer STAT1 (pY701) ELISA Kit

ab119671: PhosphoTracer STAT1 (pT701) + total GAPDH ELISA Kit

ab119678: PhosphoTracer STAT1 (pY701) + STAT3 (pY705) + STAT5 (pY694) ELISA Kit

We have several antibodies to IRAK (ab47843 and ab5522), STAT1 (ab118848 and ab64913), and IRF5 (ab33478 and ab49056)that would be suitable for use in ELISA, but none of our TRAF6 antibodies have been validated for this purpose. If you would like to test one of our antibodies in an untested species or application, I would be happy to send you more information about our testing discount program.

Abcam Scientific Support

Answered on Feb 14 2012

Question

I would like to know whether you have any flow cytometry antibodies for TRAF6 and IRAK1? Or for TRAF6 and IRAK1 protein studies in whole or peripheral blood?

Abcam community

Verified customer

Asked on Feb 13 2012

Answer

Thank you for contacting us. We currently have one antibody for each of these targets that has been validated for use in Flow Cytometry:

ab238 rabbit polyclonal to IRAK; tested for use in WB, IP, IHC-P, Flow Cyt, ICC/IF; reacts with human, mouse, and rat.

ab33915 rabbit monoclonal to TRAF6; tested for use in WB, IP, Flow Cyt, IHC-P; reacts with human, mouse, and rat.

Both of these antibodies are unconjugated, but you could use on of our EasyLink conjugation kits to directly conjugate ab238. If you are looking for an isotype control, you may be interested in ab27478.

https://www.abcam.com/EasyLink

The rabbit monoclonal ab33915 typically has a very low concentration because rabbit monoclonals have a much higher binding affinity than mouse monoclonals, but the hybridomas produce less antibody. For this reason, the concentration will be too low for use with our EasyLink conjugation kits, but it can be used with an anti-rabbit IgG secondary antibody.

I hope this helps, please let me know if you need any additional information.

Abcam Scientific Support

Answered on Feb 13 2012

Abcam guarantees this product to work in the species/application used in this Abreview.

Application

Western blot

Sample

Human Cell lysate - whole cell (HeLa Cells)

Loading amount

25 µg

Specification

HeLa Cells

Gel Running Conditions

Non-reduced Denaturing (10% gel)

Blocking step

Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C

Abcam user community

Verified customer

Submitted Feb 01 2012

Abcam guarantees this product to work in the species/application used in this Abreview.

Application

Western blot

Sample

Human Cell lysate - whole cell (Platelets)

Loading amount

20 µg

Specification

Platelets

Treatment

ADP for 30 min

Gel Running Conditions

Reduced Denaturing (4-20%)

Blocking step

Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C

Dr. Mahesh Shivananjappa

Verified customer

Submitted Nov 23 2011

Abcam guarantees this product to work in the species/application used in this Abreview.

Application

Flow Cytometry

Sample

Human Cell (Platelets)

Specification

Platelets

Preparation

Cell harvesting/tissue preparation method: PL were isolated spinning Platelet rich plasma on Histopaque
Sample buffer: PBS

Fixation

Paraformaldehyde

Permeabilization

Yes - 0.1% Triton-X100 in 2% BSA for 15min

Gating Strategy

Platelets

Abcam user community

Verified customer

Submitted Dec 21 2010

Abcam guarantees this product to work in the species/application used in this Abreview.

Application

Western blot

Sample

Mouse Cell lysate - whole cell (RAW264.7)

Loading amount

50000 cells

Specification

RAW264.7

Treatment

100 ng/ml recombinant mouse RANKL

Gel Running Conditions

Reduced Denaturing (NuPage 10% Bis-Tris)

Blocking step

BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Mar 01 2010

1-10 of 16 Abreviews or Q&A

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Anti-TRAF6 antibody [EP591Y] (ab33915)

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This product has been referenced in:

Customer reviews and Q&As

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Western blot abreview for Anti-TRAF6 antibody [EP591Y]

Western blot abreview for Anti-TRAF6 antibody [EP591Y]

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Western blot abreview for Anti-TRAF6 antibody [EP591Y]