Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-TRAF2 antibody [EPR6048] - BSA and Azide free (ab230795)

Overview

  • Product name

    Anti-TRAF2 antibody [EPR6048] - BSA and Azide free
    See all TRAF2 primary antibodies
  • Description

    Rabbit monoclonal [EPR6048] to TRAF2 - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, IP, IHC-P, Flow Cyt, ICC/IFmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide, corresponding to N terminal residues of Human TRAF2.

  • Positive control

    • PC12, MOLT4, HeLa, and 293T cell lysates; HeLa cells; Human kidney tissue.
  • General notes

    Ab230795 is the carrier-free version of ab126758. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits  for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    ab230795 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

Properties

Applications

Our Abpromise guarantee covers the use of ab230795 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 Use at an assay dependent concentration. Detects a band of approximately 53 kDa (predicted molecular weight: 55 kDa).
IP Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

Flow Cyt Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG (Low endotoxin, Azide free), is suitable for use as an isotype control with this antibody.

ICC/IF Use at an assay dependent concentration.

Target

  • Function

    Regulates activation of NF-kappa-B and JNK and plays a central role in the regulation of cell survival and apoptosis. Required for normal antibody isotype switching from IgM to IgG. Has E3 ubiquitin-protein ligase activity and promotes 'Lys-63'-linked ubiquitination of target proteins, such as BIRC3, RIPK1 and TICAM1. Is an essential constituent of several E3 ubiquitin-protein ligase complexes, where it promotes the ubiquitination of target proteins by bringing them into contact with other E3 ubiquitin ligases. Regulates BIRC2 and BIRC3 protein levels by inhibiting their autoubiquitination and subsequent degradation; this does not depend on the TRAF2 RING-type zinc finger domain.
  • 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.
    The RING-type zinc finger domain is essential for E3 ubiquitin-protein ligase activity. It is not essential for the stabilization of BIRC2, or for the ubiquitination of RIPK1 in response to TNFR1 signaling.
  • Post-translational
    modifications

    Phosphorylated at several serine residues within the first 128 amino acid residues. Phosphorylated at Thr-117 in response to signaling via TNF and TNFRSF1A. Phosphorylation at Thr-117 is required for 'Lys-63'-linked polyubiquitination, but not for 'Lys-48'-linked polyubiquitination. Phosphorylation at Thr-117 is important for interaction with IKKA and IKKB, activation of IKK and subsequent activation of NF-kappa-B.
    Undergoes both 'Lys-48'-linked and 'Lys-63'-linked polyubiquitination. Polyubiquitinated via 'Lys-63'-linked ubiquitin in response to TNF signaling; this requires prior phosphorylation at Thr-117. 'Lys-63'-linked polyubiquitination promotes TRAF2-mediated activation of NF-kappa-B. Can be polyubiquitinated at several Lys residues via 'Lys-48'-linked ubiquitin chains in response to TNF signaling, leading to proteasomal degradation. Autoubiquitinated, leading to its subsequent proteasomal degradation. Polyubiquitinated by BIRC2 and SIAH2, leading to its subsequent proteasomal degradation. Deubiquitinated by CYLD, a protease that specifically cleaves 'Lys-63'-linked polyubiquitin chains.
  • Cellular localization

    Cytoplasm.
  • Information by UniProt
  • Database links

  • Alternative names

    • E3 ubiquitin-protein ligase TRAF2 antibody
    • MGC:45012 antibody
    • OTTHUMP00000022625 antibody
    • OTTHUMP00000064745 antibody
    • TNF receptor associated factor 2 antibody
    • TNF receptor-associated factor 2 antibody
    • TNF receptor-associated protein antibody
    • TRAF 2 antibody
    • TRAF2 antibody
    • TRAF2_HUMAN antibody
    • TRAP 3 antibody
    • TRAP antibody
    • TRAP3 antibody
    • Tumor necrosis factor type 2 receptor associated protein 3 antibody
    • Tumor necrosis factor type 2 receptor-associated protein 3 antibody
    see all

Images

  • This WB data was generated using the same anti-TRAF2 antibody clone, EPR6048, in a different buffer formulation (cat# ab126758).

    Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: TRAF2 knockout HAP1 cell lysate (20 µg) 
    Lane 3: Human skeletal muscle lysate (20 µg)
    Lane 4: U2OS cell lysate (20 µg) 
    Lanes 1 - 4: Merged signal (red and green). Green - ab126758 observed at 55 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab126758 was shown to specifically react with TRAF2 when TRAF2 knockout samples were used. Wild-type and TRAF2 knockout samples were subjected to SDS-PAGE. ab126758 and ab8245 (loading control to GAPDH) were diluted 1/1000 and 1/2000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.

  • ab126758 (purified) at 1/40 immunoprecipitating TRAF2 in HEK293 (Lane 1 and 2). Lane 3 - PBS. For western blotting a HRP-conjugated anti-rabbit IgG specific to the non-reduced form of IgG was used as the secondary antibody (1/1500). Blocking buffer and concentration: 5% NFDM/TBST. Diluting buffer and concentration: 5% NFDM /TBST.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab126758).

  • Overlay histogram showing HeLa cells fixed in 2% PFA and stained with purified ab126758 at a dilution of 1 in 120 (red line). The secondary antibody used was FITC goat anti-rabbit at a dilution of 1 in 150. Rabbit monoclonal IgG was used as an isotype control (black) and cells without antibody were used as a negative control (blue).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab126758).

  • Immunofluorescence staining of HeLa cells with purified ab126758 at a working dilution of 1 in 100, counter-stained with DAPI. Tubulin was stained with mouse anti-tubulin at a dilution of 1/1000 (ab7291) and Alexa Fluor® 594 goat anti-mouse at a dilution of 1/500 (ab150120) . The secondary antibody was ab150077 Alexa Fluor® 488 goat anti rabbit, used at a dilution of 1 in 500. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative controls are shown in the bottom middle and right hand panels - for the first negative control, purified ab126758 was used at a dilution of 1/200 followed by an Alexa Fluor® 555 goat anti-mouse antibody at a dilution of 1/500 and for the second negative control mouse primary antibody (ab7291) and anti-rabbit secondary antibody (ab15007) were used.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab126758).

  • Overlay histogram showing HeLa cells stained with unpurified ab126758 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (unpurified ab126758, 1/100 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab126758).

  • Unpurified ab126758, at 1/50 dilution, staining TRAF2 in paraffin-embedded Human kidney tissue, by Immunohistochemistry.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab126758).

  • Unpurified ab126758, at 1/100 dilution, staining TRAF2 in HeLa cells, by Immunofluorescence.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab126758).

  • Equilibrium disassociation constant (KD)
    Learn more about KD

    Click here to learn more about KD

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab126758).

  • This IHC data was generated using the same anti-TRAF2 antibody clone, EPR6048, in a different buffer formulation (cat# ab126758).

    Immunohistochemical staining of paraffin embedded human cervical cancer with purified ab126758 at a working dilution of 1/100. The secondary antibody used is ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L), at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

References

ab230795 has not yet been referenced specifically in any publications.

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