Anti-DNA PKcs (phospho S2056) antibody - ChIP Grade (ab18192)
Overview
- Product nameAnti-DNA PKcs (phospho S2056) antibody - ChIP GradeSee all DNA PKcs primary antibodies ...
- DescriptionRabbit polyclonal to DNA PKcs (phospho S2056) - ChIP Grade
- SpecificityThis antibody specifically recognizes a band at ~460 kDa in HeLa cells that have been treated with ionizing radiation, that is not detected in untreated cells. This band can also be competed away by the immunizing modified peptide, but not the unmodified peptide containing the same amino acid sequence. All batches of this antibody are screened in ELISA and show high titres against the immunising peptide. Reactivity with the unmodified DNA PKcs peptide is minimal.
- Tested applicationsFlow Cyt, IHC-P, WB, ChIP, ICC/IF more details
- Species reactivityReacts with: Human
Does not react with
Mouse - Immunogen
Synthetic peptide conjugated to KLH derived from within residues 2050 - 2150 of Human DNA PKcs.
(Peptide available as ab20406.)
- Positive controlThis antibody gave a positive signal in Hela cells which have been treated with 20-40 mU/ml bleomycin for 48hrs.
Properties
- FormLiquid
- Storage instructionsStore at +4°C short term (1-2 weeks). Aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
- Storage bufferPreservative: 0.02% Sodium Azide
Constituents: 1% BSA, PBS, pH 7.4 -
Concentration information loading... - PurityImmunogen affinity purified
- Clonality Polyclonal
- IsotypeIgG
- Research Areas
Applications
Our Abpromise guarantee covers the use of ab18192 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Notes |
|---|---|
| Flow Cyt | Flow Cyt: Use at an assay dependent dilution. |
| IHC-P | IHC-P: Use at an assay dependent concentration. |
| WB | WB: Use a concentration of 1 µg/ml. Detects a band of approximately 460 kDa (predicted molecular weight: 460 kDa).Can be blocked with DNA PKcs peptide - phospho S2056 (ab20406). |
| ChIP | ChIP: Use at an assay dependent dilution. PubMed: 20714222 |
| ICC/IF | ICC/IF: Use a concentration of 1 µg/ml. |
Target
- FunctionSerine/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 similaritiesBelongs 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
modificationsPhosphorylated 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 localizationNucleus.
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Database links
- Entrez Gene: 5591 Human
- Omim: 600899 Human
- SwissProt: P78527 Human
- Unigene: 491682 Human
Target information above from: UniProt accession
P78527
The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010)
.
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Alternative names
- DNA dependent protein kinase catalytic subunit antibodyDNA PK catalytic subunit antibodyDNA-dependent protein kinase catalytic subunit antibody
- DNA-PK catalytic subunit antibodyDNA-PKcs antibodyDNAPK antibodyDNAPK catalytic subunit antibodyDNPK 1 antibodyDNPK1 antibodyDNPK1 antibodyHyper radiosensitivity of murine scid mutation, complementing 1 antibodyHYRC 1 antibodyHYRC antibodyHYRC1 antibodyHYRC1 antibodyp350 antibodyp460 antibodyPKRDC antibodyPRKDC antibodyPRKDC_HUMAN antibodyProtein Kinase DNA Activated Catalytic Polypeptide antibodyProtein Kinase DNA Activated Catalytic Polypeptide antibodyXRCC 7 antibodyXRCC7 antibodyXRCC7 antibody
see all
Anti-DNA PKcs (phospho S2056) antibody - ChIP Grade images
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Western blot - DNA PKcs (phospho S2056) antibody (ab18192)All lanes : Anti-DNA PKcs (phospho S2056) antibody - ChIP Grade (ab18192) at 1 µg/ml
Lane 1 : HeLa Gamma Irradiated Whole Cell Lysate Pack (ab13823) at 20 µg
Lane 2 : 20ug of untreated HeLa cell extract
Lane 3 : HeLa Gamma Irradiated Whole Cell Lysate Pack (ab13823) at 20 µg withDNA PKcs peptide - phospho S2056 (ab20406) at 1 µg/ml
Lane 4 : 20ug of untreated HeLa cell extract withDNA PKcs peptide - phospho S2056 (ab20406) at 1 µg/ml
Lane 5 : HeLa Gamma Irradiated Whole Cell Lysate Pack (ab13823) at 20 µg withDNA PKcs peptide (ab20407) at 1 µg/ml
Lane 6 : 20ug of untreated HeLa cell extract withDNA PKcs peptide (ab20407) at 1 µg/ml
Secondary
Alexa Fluor Goat polyclonal to Rabbit IgG (700) at 1/10000 dilution
developed using the ECL technique
Performed under reducing conditions.
Predicted band size : 460 kDa
Observed band size : 460 kDa
Additional bands at : 270 kDa (possible cleavage fragment,cross reactivity). -
Immunocytochemistry/ Immunofluorescence - DNA PKcs (phospho S2056) antibody (ab18192)ICC/IF image of ab18192 stained HepG2 cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal got serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab18192, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 4% PFA fixed (10 min)HeLa, Hek293 and MCF7 cells at 1µg/ml, and in 100% methanol fixed (5 min) HeLa, Hek293 and HepG2 cells at 5µg/ml. -
Western blot - DNA PKcs (phospho S2056) antibody (ab18192)All lanes : Anti-DNA PKcs (phospho S2056) antibody - ChIP Grade (ab18192) at 1 µg/ml
Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
Lane 2 : Hela Whole Cell Lysate - Bleomycin Treated (20U/ml)
Lane 3 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate withDNA PKcs peptide - phospho S2056 (ab20406) at 1 µg/ml
Lane 4 : Hela Whole Cell Lysate - Bleomycin Treated (20U/ml) withDNA PKcs peptide - phospho S2056 (ab20406) at 1 µg/ml
Lane 5 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate withDNA PKcs peptide (ab20407) at 1 µg/ml
Lane 6 : Hela Whole Cell Lysate - Bleomycin Treated (20U/ml) withDNA PKcs peptide (ab20407) at 1 µg/ml
Lysates/proteins at 20 µg per lane.
Secondary
Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
Performed under reducing conditions.
Predicted band size : 460 kDa
Observed band size : 460 kDa
Additional bands at : 117 kDa,150 kDa,270 kDa,50 kDa. We are unsure as to the identity of these extra bands.
Exposure time : 15 minutes -
Flow Cytometry - Anti-DNA PKcs (phospho S2056) antibody - ChIP Grade (ab18192)This image is courtesy of an Anonymous Abreview.Overlay histogram showing Jurkat cells stained with ab18192 (blue line). The cells were prepared as follows: Spin down, wash in FACS buffer 1x, fix, wash 2x, and stain with primary antibody overnight. Buffer was was 0.1% sodium azide with FBS in PBS. Cells fixed with paraformaldehyde and then permeabilized with Triton X-100 and NP40. Cell population was gated by isolating cell population from plot of SSC-A / FSA-A. The cells were then incubated the antibody at 1/100 for 24 hours. The secondary antibody used was FITC-conjugated Goat anti-Rabbit polyclonal at 1/100 dilution. Isotype control antibody (red line) was used under the same conditions. -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-DNA PKcs (phospho S2056) antibody - ChIP Grade (ab18192)This image is courtesy of an anonymous Abreviewab18192 staining DNA PKcs in Human aorta tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and blocked with 100% blocking agent for 20 minutes; antigen retrieval was enzymatic using Proteinase K. Samples were incubated with primary antibody (1/350 in diluent) for 15 hours at 4°C. A biotinylated Goat anti-rabbit polyclonal (1/800) was used as the secondary antibody. -
Immunocytochemistry/ Immunofluorescence - Anti-DNA PKcs (phospho S2056) antibody - ChIP Grade (ab18192)Image from Furusawa Y et al., PLoS One. 2012;7(1):e29012. Epub 2012 Jan 3. Fig 2.; doi:10.1371/journal.pone.0029012; January 3, 2012, PLoS ONE 7(1): e29012.ab18192 staining DNA PKcs (phospho S2056) in leukaemia cell lines by Immunocytochemistry/ Immunofluorescence.
Cells were either untreated (control) or treated with ionizing radiation (IR) or ultrasound (US). Following treatment, cells were fixed with paraformaldehyde and permabilized/blocked with 2% BSA/0.05% Triton X-100/Tris-buffered saline. Samples were incubated with primary antibody (1/600 in diluent) for 2 hours. An AlexaFluor®555-conjugated anti-rabbit IgG (1/400) was used as the secondary antibody.
References for Anti-DNA PKcs (phospho S2056) antibody - ChIP Grade (ab18192)
This product has been referenced in:
- Valerie NC et al. Inhibition of T-type calcium channels disrupts Akt signaling and promotes apoptosis in glioblastoma cells. Biochem Pharmacol 85:888-97 (2013). Read more (PubMed: 23287412) »
- Meek K et al. N-terminal constraint activates the catalytic subunit of the DNA-dependent protein kinase in the absence of DNA or Ku. Nucleic Acids Res 40:2964-73 (2012). WB . Read more (PubMed: 22167471) »
