Validated using a knockout cell line
Recombinant
RabMAb

Anti-53BP1 antibody [EPR2172(2)] (ab175933)

Overview

  • Product name
    Anti-53BP1 antibody [EPR2172(2)]
    See all 53BP1 primary antibodies
  • Description
    Rabbit monoclonal [EPR2172(2)] to 53BP1
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WB, IHC-P, ICC/IF, Flow Cytmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) within Human 53BP1 aa 1-100 (Cysteine residue). The exact sequence is proprietary.
    Database link: Q12888

  • Positive control
    • WB: HepG2 and HeLa cell lysate and human fetal heart and fetal brain tissue lysates, mouse heart and rat heart tissue lysates. IHC-P: human colon, liver carcinoma and tonsil, mouse and rat liver tissues. ICC/IF: HepG2 cells.
  • 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

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab175933 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 1/1000 - 1/5000. Detects a band of approximately 450 kDa (predicted molecular weight: 214 kDa).
IHC-P 1/60 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

ICC/IF 1/100 - 1/250.
Flow Cyt Use at an assay dependent concentration.

Target

  • Function
    May have a role in checkpoint signaling during mitosis. Enhances TP53-mediated transcriptional activation. Plays a role in the response to DNA damage.
  • Involvement in disease
    Note=A chromosomal aberration involving TP53BP1 is found in a form of myeloproliferative disorder chronic with eosinophilia. Translocation t(5;15)(q33;q22) with PDGFRB creating a TP53BP1-PDGFRB fusion protein.
  • Sequence similarities
    Contains 2 BRCT domains.
  • Post-translational
    modifications
    Asymmetrically dimethylated on Arg residues by PRMT1. Methylation is required for DNA binding.
    Phosphorylated at basal level in the absence of DNA damage. Hyper-phosphorylated in an ATM-dependent manner in response to DNA damage induced by ionizing radiation. Hyper-phosphorylated in an ATR-dependent manner in response to DNA damage induced by UV irradiation.
  • Cellular localization
    Nucleus. Chromosome > centromere > kinetochore. Associated with kinetochores. Both nuclear and cytoplasmic in some cells. Recruited to sites of DNA damage, such as double stand breaks. Methylation of histone H4 at 'Lys-20' is required for efficient localization to double strand breaks.
  • Information by UniProt
  • Database links
  • Alternative names
    • 53 BP1 antibody
    • 53BP1 antibody
    • FLJ41424 antibody
    • MGC138366 antibody
    • p202 antibody
    • p53 binding protein 1 antibody
    • p53 BP1 antibody
    • p53-binding protein 1 antibody
    • p53BP1 antibody
    • TP53 BP1 antibody
    • TP53B_HUMAN antibody
    • Tp53bp1 antibody
    • TRP53 BP1 antibody
    • Tumor protein 53 binding protein 1 antibody
    • Tumor protein p53 binding protein 1 antibody
    • Tumor suppressor p53 binding protein 1 antibody
    • Tumor suppressor p53-binding protein 1 antibody
    see all

Images

  • Immunocytochemistry/Immunofluorescence analysis of HepG2 cells labelling 53BP1 with purified ab175933 at 1/200. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) were also used.

    Control 1: primary antibody (1/200) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000).

    Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000).

  • Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: 53BP1 knockout HAP1 cell lysate (20 µg)
    Lane 3: HeLa cell lysate (40 µg)
    Lane 4: HepG2 cell lysate (40 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab175933 observed at 350 kDa. Red - loading control, ab18058, observed at 124 kDa.
    ab175933 was shown to specifically react with 53BP1 when 53BP1 knockout samples were used. Wild-type and 53BP1 knockout samples were subjected to SDS-PAGE. ab175933 and ab18058 (loading control to Vinculin) were diluted 1/1000 and 1/10000 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 hour at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat liver tissue labelling 53BP1 with purified ab175933 at a dilution of 1/60. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • ab175933 staining 53BP1 in the human cell line HepG2 (human hepatocellular carcinoma) by flow cytometry. Cells were fixed with 4% paraformaldehyde and the sample was incubated with the primary antibody at a dilution of 1/30. A goat anti rabbit IgG (Alexa Fluor® 488) at a dilution of 1/2000 was used as the secondary antibody.

    Isoytype control: Rabbit monoclonal IgG (Black)

    Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue)

  • Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: 53BP1 knockout HAP1 cell lysate (20 µg)
    Lane 3: HeLa cell lysate (40 µg)
    Lane 4: HepG2 cell lysate (40 µg)

    Lanes 1 - 4: Merged signal (red and green).

    Green - Target observed at 350 kDa. Red - loading control, ab18058, observed at 124 kDa.

    This western blot image is a comparison between ab175933 and a competitor's top cited rabbit polyclonal antibody.

  • All lanes : Anti-53BP1 antibody [EPR2172(2)] (ab175933) at 1/1000 dilution (purified)

    Lane 1 : Mouse heart tissue lysate
    Lane 2 : Mouse brain tissue lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 214 kDa
    Observed band size: 450 kDa (why is the actual band size different from the predicted?)



    Blocking and dilution buffer: 5% NFDM /TBST.

  • Anti-53BP1 antibody [EPR2172(2)] (ab175933) at 20 µg (purified) + Rat heart tissue lysate at 20 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 214 kDa
    Observed band size: 450 kDa (why is the actual band size different from the predicted?)



    Blocking and dilution buffer: 5% NFDM /TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse liver tissue labelling 53BP1 with purified ab175933 at a dilution of 1/60. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • All lanes : Anti-53BP1 antibody [EPR2172(2)] (ab175933) at 1/5000 dilution (purified)

    Lane 1 : Human fetal heart tissue lysate
    Lane 2 : HeLa 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

    Predicted band size: 214 kDa
    Observed band size: 450 kDa (why is the actual band size different from the predicted?)



    Blocking and dilution buffer: 5% NFDM /TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human liver carcinoma tissue labelling 53BP1 with purified ab175933 at a dilution of 1/60. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

  • All lanes : Anti-53BP1 antibody [EPR2172(2)] (ab175933) at 1/1000 dilution (unpurified)

    Lane 1 : HepG2 cell lysate
    Lane 2 : Human fetal brain lysate
    Lane 3 : Human fetal heart lysate

    Lysates/proteins at 1/10 dilution per lane.

    Predicted band size: 214 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human tonsil tissue labelling 53BP1 with unpurified ab175933 at a dilution of 1/100.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colon tissue labelling 53BP1 with unpurified ab175933 at a dilution of 1/100.

  • Immunocytochemistry/Immunofluorescence analysis of HepG2 cells labelling 53BP1 with unpurified ab175933 at a dilution of 1/100.

References

This product has been referenced in:

See all 3 Publications for this product

Customer reviews and Q&As

Application
Immunocytochemistry
Sample
Human Cell (HeLa)
Permeabilization
Yes - 0.5% TritonX100 in PBS
Specification
HeLa
Fixative
Paraformaldehyde
Username

Dr. Kirk Mcmanus

Verified customer

Submitted Apr 22 2015

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HeLa)
Specification
HeLa
Permeabilization
Yes - 0.5% Triton X100 in PBS
Fixative
Paraformaldehyde
Username

Dr. Kirk Mcmanus

Verified customer

Submitted Jul 10 2014

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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