Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-DNA PKcs antibody [EPR392] (ab133516)

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Western blot - Anti-DNA PKcs antibody [EPR392] (ab133516)
  • Western blot - Anti-DNA PKcs antibody [EPR392] (ab133516)
  • Immunocytochemistry/ Immunofluorescence - Anti-DNA PKcs antibody [EPR392] (ab133516)
  • OI-RD Scanning - Anti-DNA PKcs antibody [EPR392] (ab133516)
  • Western blot - Anti-DNA PKcs antibody [EPR392] (ab133516)
  • Anti-DNA PKcs antibody [EPR392] (ab133516)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR392] to DNA PKcs
  • Suitable for: WB, ICC/IF
  • Knockout validated
  • Reacts with: Human

Overview

  • Product name

    Anti-DNA PKcs antibody [EPR392]
    See all DNA PKcs primary antibodies

  • Description

    Rabbit monoclonal [EPR392] to DNA PKcs

  • Host species

    Rabbit

  • Tested applications

    Suitable for: WB, ICC/IFmore details
    Unsuitable for: Flow Cyt,IHC-P or IP

  • Species reactivity

    Reacts with: Human

  • Immunogen

    Synthetic peptide within Human DNA PKcs aa 50-150. The exact sequence is proprietary.
    Database link: P78527

  • Positive control

    • WB: K562, Molt4, MCF7, SH-SY5Y, 293T and PC3 cell lysates. ICC/IF: K-562 cells

  • General notes

    Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.

     

    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
    For more information see here.

    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.

    Reproducibility is key to advancing scientific discovery and accelerating scientists’ next breakthrough.

    Abcam is leading the way with our range of recombinant antibodies, knockout-validated antibodies and knockout cell lines, all of which support improved reproducibility.

    We are also planning to innovate the way in which we present recommended applications and species on our product datasheets, so that only applications & species that have been tested in our own labs, our suppliers or by selected trusted collaborators are covered by our Abpromise™ guarantee.

    In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.

    We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.

    Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.

    Please check that this product meets your needs before purchasing. If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, as well as customer reviews and Q&As.

Properties

Applications

Our Abpromise guarantee covers the use of ab133516 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 - 1/10000. Detects a band of approximately 460 kDa (predicted molecular weight: 469 kDa).
ICC/IF 1/200.

For unpurified use at 1/500 - 1/1000.
  • Application notes
    Is unsuitable for Flow Cyt,IHC-P or IP.

    • Target

    • Function

      Serine/threonine-protein kinase that acts as a molecular sensor for DNA damage. Involved in DNA nonhomologous end joining (NHEJ) required for double-strand break (DSB) repair and V(D)J recombination. Must be bound to DNA to express its catalytic properties. Promotes processing of hairpin DNA structures in V(D)J recombination by activation of the hairpin endonuclease artemis (DCLRE1C). The assembly of the DNA-PK complex at DNA ends is also required for the NHEJ ligation step. Required to protect and align broken ends of DNA. May also act as a scaffold protein to aid the localization of DNA repair proteins to the site of damage. Found at the ends of chromosomes, suggesting a further role in the maintenance of telomeric stability and the prevention of chromosomal end fusion. Also involved in modulation of transcription. Recognizes the substrate consensus sequence [ST]-Q. Phosphorylates 'Ser-139' of histone variant H2AX/H2AFX, thereby regulating DNA damage response mechanism. Phosphorylates DCLRE1C, c-Abl/ABL1, histone H1, HSPCA, c-jun/JUN, p53/TP53, PARP1, POU2F1, DHX9, SRF, XRCC1, XRCC1, XRCC4, XRCC5, XRCC6, WRN, MYC and RFA2. Can phosphorylate C1D not only in the presence of linear DNA but also in the presence of supercoiled DNA. Ability to phosphorylate p53/TP53 in the presence of supercoiled DNA is dependent on C1D.

    • Sequence similarities

      Belongs to the PI3/PI4-kinase family.
      Contains 1 FAT domain.
      Contains 1 FATC domain.
      Contains 2 HEAT repeats.
      Contains 1 PI3K/PI4K domain.
      Contains 3 TPR repeats.

    • Post-translational
      modifications

      Phosphorylated upon DNA damage, probably by ATM or ATR. Autophosphorylated on Thr-2609, Thr-2638 and Thr-2647. Thr-2609 is a DNA damage-inducible phosphorylation site (inducible with ionizing radiation, IR). Autophosphorylation induces a conformational change that leads to remodeling of the DNA-PK complex, requisite for efficient end processing and DNA repair.
      S-nitrosylated by GAPDH.

    • Cellular localization

      Nucleus.
    • Information by UniProt

    • Database links

      • Alternative names

        • DNA dependent protein kinase catalytic subunit antibody
        • DNA PK catalytic subunit antibody
        • DNA-dependent protein kinase catalytic subunit antibody
        • DNA-PK catalytic subunit antibody
        • DNA-PKcs antibody
        • DNAPK antibody
        • DNAPK catalytic subunit antibody
        • DNPK 1 antibody
        • DNPK1 antibody
        • Hyper radiosensitivity of murine scid mutation, complementing 1 antibody
        • Hyperradiosensitivity complementing 1, mouse, homolog of 1 antibody
        • HYRC 1 antibody
        • HYRC antibody
        • HYRC1 antibody
        • IMD26 antibody
        • p350 antibody
        • p460 antibody
        • PKRDC antibody
        • PRKDC antibody
        • PRKDC_HUMAN antibody
        • Protein Kinase DNA Activated Catalytic Polypeptide antibody
        • XRCC 7 antibody
        • XRCC7 antibody
        see all

      Images

      • Western blot - Anti-DNA PKcs antibody [EPR392] (ab133516)
        Western blot - Anti-DNA PKcs antibody [EPR392] (ab133516)

        Lane 1: Wild type HAP1 whole cell lysate (40 µg)
        Lane 2: DNA PKcs knockout HAP1 whole cell lysate (40 µg)
        Lane 3: K562 whole cell lysate (20 µg)
        Lane 4: SHSY5Y whole cell lysate (20 µg)

        Lanes 1 - 4: Merged signal (red and green). Green - ab133516 observed at 460 kDa. Red - loading control, ab18058, observed at 130 kDa.

        ab133516 was shown to specifically react with DNA PKcs when DNA PKcs knockout samples were used. Wild-type and DNA PKcs knockout samples were subjected to SDS-PAGE. Ab133516 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 1000 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.

         

      • Western blot - Anti-DNA PKcs antibody [EPR392] (ab133516)
        Western blot - Anti-DNA PKcs antibody [EPR392] (ab133516)
        All lanes : Anti-DNA PKcs antibody [EPR392] (ab133516) at 1/5000 dilution (Purified)

        Lane 1 : K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysates
        Lane 2 : SH-SY5Y (Human neuroblastoma epithelial cell) 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

        Predicted band size: 469 kDa
        Observed band size: 460 kDa
        why is the actual band size different from the predicted?

      • Immunocytochemistry/ Immunofluorescence - Anti-DNA PKcs antibody [EPR392] (ab133516)
        Immunocytochemistry/ Immunofluorescence - Anti-DNA PKcs antibody [EPR392] (ab133516)
        Immunocytochemistry/ Immunofluorescence analysis of K-562 (Human chronic myelogenous leukemia lymphoblast) cells labeling DNA PKcs with purified ab133516 at 1:200 dilution (9.3 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1:200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1:1000 (2 µg/ml) dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
      • OI-RD Scanning - Anti-DNA PKcs antibody [EPR392] (ab133516)
        OI-RD Scanning - Anti-DNA PKcs antibody [EPR392] (ab133516)
        Equilibrium disassociation constant (KD)
        Learn more about KD

        Click here to learn more about KD
      • Western blot - Anti-DNA PKcs antibody [EPR392] (ab133516)
        Western blot - Anti-DNA PKcs antibody [EPR392] (ab133516)
        All lanes : Anti-DNA PKcs antibody [EPR392] (ab133516) at 1/1000 dilution

        Lane 1 : K562 cell lysate
        Lane 2 : Molt4 cell lysate
        Lane 3 : MCF7 cell lysate
        Lane 4 : SH-SY5Y cell lysate
        Lane 5 : 293T cell lysate
        Lane 6 : PC3 cell lysate

        Lysates/proteins at 10 µg per lane.

        Secondary
        All lanes : Goat-anti-rabbit HRP at 1/2000 dilution

        Predicted band size: 469 kDa
        Observed band size: 460 kDa why is the actual band size different from the predicted?

      • Anti-DNA PKcs antibody [EPR392] (ab133516)
        Anti-DNA PKcs antibody [EPR392] (ab133516)

      References (1)

      Publishing research using ab133516? Please let us know so that we can cite the reference in this datasheet.

      ab133516 has been referenced in 1 publication.

      • Fujiwara Y  et al. A Nucleolar Stress-Specific p53-miR-101 Molecular Circuit Functions as an Intrinsic Tumor-Suppressor Network. EBioMedicine 33:33-48 (2018). PubMed: 30049386

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