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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)

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Western blot - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
  • Western blot - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
  • Western blot - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
  • Immunocytochemistry/ Immunofluorescence - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
  • Immunocytochemistry/ Immunofluorescence - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
  • Flow Cytometry (Intracellular) - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
  • Immunoprecipitation - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
  • ChIP - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
  • Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR20100] to ATM - ChIP Grade
  • Suitable for: Flow Cyt (Intra), ICC/IF, IP, ChIP, WB
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Carrier Free

Overview

  • Product name

    Anti-ATM antibody [EPR20100] - ChIP Grade
    See all ATM primary antibodies

  • Description

    Rabbit monoclonal [EPR20100] to ATM - ChIP Grade

  • Host species

    Rabbit

  • Tested applications

    Suitable for: Flow Cyt (Intra), ICC/IF, IP, ChIP, WBmore details

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: Human testis lysate; HeLa, PC-12, RAW 264.7, 293 and SH-SY5Y whole cell lysates. ICC/IF: SH-SY5Y and HeLa cells. Flow Cyt (intra): HEK-293 cells. IP: HEK-293 whole cell lysate. ChIP: Chromatin prepared from HCT 116 cells treated with 1mM Hydroxyurea for 16h.

  • General notes

    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.

Properties

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab201022 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
Flow Cyt (Intra)
1/800.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
ICC/IF
1/500.
IP
1/40.
ChIP
Use 2 µg for 25 µg of chromatin.
WB
1/1000. Detects a band of approximately 351 kDa (predicted molecular weight: 351 kDa).
Notes
Flow Cyt (Intra)
1/800.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
ICC/IF
1/500.
IP
1/40.
ChIP
Use 2 µg for 25 µg of chromatin.
WB
1/1000. Detects a band of approximately 351 kDa (predicted molecular weight: 351 kDa).

Target

  • Function

    Serine/threonine protein kinase which activates checkpoint signaling upon double strand breaks (DSBs), apoptosis and genotoxic stresses such as ionizing ultraviolet A light (UVA), thereby acting as a DNA damage sensor. Recognizes the substrate consensus sequence [ST]-Q. Phosphorylates 'Ser-139' of histone variant H2AX/H2AFX at double strand breaks (DSBs), thereby regulating DNA damage response mechanism. Also plays a role in pre-B cell allelic exclusion, a process leading to expression of a single immunoglobulin heavy chain allele to enforce clonality and monospecific recognition by the B-cell antigen receptor (BCR) expressed on individual B lymphocytes. After the introduction of DNA breaks by the RAG complex on one immunoglobulin allele, acts by mediating a repositioning of the second allele to pericentromeric heterochromatin, preventing accessibility to the RAG complex and recombination of the second allele. Also involved in signal transduction and cell cycle control. May function as a tumor suppressor. Necessary for activation of ABL1 and SAPK. Phosphorylates p53/TP53, FANCD2, NFKBIA, BRCA1, CTIP, nibrin (NBN), TERF1, RAD9 and DCLRE1C. May play a role in vesicle and/or protein transport. Could play a role in T-cell development, gonad and neurological function. Plays a role in replication-dependent histone mRNA degradation. Binds DNA ends.

  • Tissue specificity

    Found in pancreas, kidney, skeletal muscle, liver, lung, placenta, brain, heart, spleen, thymus, testis, ovary, small intestine, colon and leukocytes.

  • Involvement in disease

    Defects in ATM are the cause of ataxia telangiectasia (AT) [MIM:208900]; also known as Louis-Bar syndrome, which includes four complementation groups: A, C, D and E. This rare recessive disorder is characterized by progressive cerebellar ataxia, dilation of the blood vessels in the conjunctiva and eyeballs, immunodeficiency, growth retardation and sexual immaturity. AT patients have a strong predisposition to cancer; about 30% of patients develop tumors, particularly lymphomas and leukemias. Cells from affected individuals are highly sensitive to damage by ionizing radiation and resistant to inhibition of DNA synthesis following irradiation.
    Note=Defects in ATM contribute to T-cell acute lymphoblastic leukemia (TALL) and T-prolymphocytic leukemia (TPLL). TPLL is characterized by a high white blood cell count, with a predominance of prolymphocytes, marked splenomegaly, lymphadenopathy, skin lesions and serous effusion. The clinical course is highly aggressive, with poor response to chemotherapy and short survival time. TPLL occurs both in adults as a sporadic disease and in younger AT patients.
    Note=Defects in ATM contribute to B-cell non-Hodgkin lymphomas (BNHL), including mantle cell lymphoma (MCL).
    Note=Defects in ATM contribute to B-cell chronic lymphocytic leukemia (BCLL). BCLL is the commonest form of leukemia in the elderly. It is characterized by the accumulation of mature CD5+ B lymphocytes, lymphadenopathy, immunodeficiency and bone marrow failure.

  • Sequence similarities

    Belongs to the PI3/PI4-kinase family. ATM subfamily.
    Contains 1 FAT domain.
    Contains 1 FATC domain.
    Contains 1 PI3K/PI4K domain.

  • Domain

    The FATC domain is required for interaction with KAT5.

  • Post-translational
    modifications

    Phosphorylated by NUAK1/ARK5. Autophosphorylation on Ser-367, Ser-1893, Ser-1981 correlates with DNA damage-mediated activation of the kinase.
    Acetylation, on DNA damage, is required for activation of the kinase activity, dimer-monomer transition, and subsequent autophosphorylation on Ser-1981. Acetylated in vitro by KAT5/TIP60.

  • Cellular localization

    Nucleus. Cytoplasmic vesicle. Primarily nuclear. Found also in endocytic vesicles in association with beta-adaptin.
  • Information by UniProt

  • Database links

    • Alternative names

      • A-T mutated antibody
      • A-T mutated homolog antibody
      • AT mutated antibody
      • AT1 antibody
      • ATA antibody
      • Ataxia telangiectasia mutated antibody
      • Ataxia telangiectasia mutated gene antibody
      • Ataxia telangiectasia mutated homolog (human) antibody
      • Ataxia telangiectasia mutated homolog antibody
      • ATC antibody
      • ATD antibody
      • ATDC antibody
      • ATE antibody
      • ATM antibody
      • ATM serine/threonine kinase antibody
      • ATM_HUMAN antibody
      • DKFZp781A0353 antibody
      • MGC74674 antibody
      • OTTHUMP00000232981 antibody
      • Serine protein kinase ATM antibody
      • Serine-protein kinase ATM antibody
      • Serine/threonine-protein kinase ATM antibody
      • Tefu antibody
      • TEL1 antibody
      • TEL1, telomere maintenance 1, homolog antibody
      • TELO1 antibody
      • Telomere fusion protein antibody
      see all

    Images

    • Western blot - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      Western blot - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      All lanes : Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022) at 1/1000 dilution

      Lane 1 : Wild-type A549 cell lysate
      Lane 2 : ATM knockout A549 cell lysate
      Lane 3 : HEK-293 cell lysate
      Lane 4 : U-2 OS cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 351 kDa
      Observed band size: 350 kDa why is the actual band size different from the predicted?



      False colour image of Western blot: Anti-ATM antibody [EPR20100] - ChIP Grade staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab201022 was shown to bind specifically to ATM. A band was observed at 350 kDa in wild-type A549 cell lysates with no signal observed at this size in ATM knockout cell line ab276095 (knockout cell lysate ab283834). To generate this image, wild-type and ATM knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5% milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

    • Western blot - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      Western blot - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022) at 1/5000 dilution + HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 10 µg

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

      Predicted band size: 351 kDa
      Observed band size: 351 kDa


      Exposure time: 10 seconds


      Blocking/Dilution buffer: 5% NFDM/TBST.

       

    • Western blot - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      Western blot - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      All lanes : Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022) at 1/1000 dilution

      Lane 1 : Human testis lysate
      Lane 2 : PC-12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate
      Lane 3 : RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate
      Lane 4 : 293 (Human epithelial cell line from embryonic kidney) whole cell lysate
      Lane 5 : SH-SY5Y (Human neuroblastoma cell line from bone marrow) whole cell 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: 351 kDa
      Observed band size: 351 kDa



      Blocking/Dilution buffer: 5% NFDM/TBST.

      Exposure time: Lane 1 and 2: 3 minutes; Lane 3: 30 seconds; Lane 4: 5 seconds; Lane 5: 1 second

    • Immunocytochemistry/ Immunofluorescence - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      Immunocytochemistry/ Immunofluorescence - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)

      Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling ATM with ab201022 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear and weak cytoplasmic staining on HeLa cell line.

      The nuclear counter stain is DAPI (blue). Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)) at 1/200 dilution (red).

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) at 1/1000 dilution.

    • Immunocytochemistry/ Immunofluorescence - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      Immunocytochemistry/ Immunofluorescence - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)

      Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized SH-SY5Y (Human neuroblastoma cell line from bone marrow) cells labeling ATM with ab201022 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear and weak cytoplasmic staining on SH-SY5Y cell line.

      The nuclear counter stain is DAPI (blue). Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)) at 1/200 dilution (red).

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) at 1/1000 dilution.

    • Flow Cytometry (Intracellular) - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      Flow Cytometry (Intracellular) - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)

      Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed HEK-293 (Human epithelial cell line from embryonic kidney) cells labeling ATM with ab201022 at 1/800 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluorr® 488) at 1/2000 dilution was used as the secondary antibody. 

       

       

    • Immunoprecipitation - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      Immunoprecipitation - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)

      ATM was immunoprecipitated from 0.35 mg of HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate with ab201022 at 1/40 dilution. Western blot was performed from the immunoprecipitate using ab201022 at 1/1000 dilution. VeriBlot for IP Detection Reaction (HRP) (ab131366), was used for detection at 1/10000 dilution.

      Lane 1: HEK-293 whole cell lysate, 10 μg (Input).

      Lane 2: ab201022 IP in HEK-293 whole cell lysate.

      Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab201022 in HEK-293 whole cell lysate.

      Blocking and dilution buffer and concentration: 5% NFDM/TBST.

      Exposure time: 3 minutes.

      ATM cleavage has been documented previously and the fragment pattern is consistent with what has been described in the literature PMID:16849690

    • ChIP - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      ChIP - Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)

      Chromatin was prepared from HCT 116 (Human colorectal carcinoma cell line) cells treated with 1mM Hydroxyurea for 16h and non-treated according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 2µg of ab201022 (blue), and 20µl of Anti rabbit IgG sepharose beads. 2μg of rabbit normal IgG was added to the beads control (yellow).  The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).

    • Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)
      Anti-ATM antibody [EPR20100] - ChIP Grade (ab201022)

    References (2)

    Publishing research using ab201022? Please let us know so that we can cite the reference in this datasheet.

    ab201022 has been referenced in 2 publications.

    • Hu S  et al. LINCS gene expression signature analysis revealed bosutinib as a radiosensitizer of breast cancer cells by targeting eIF4G1. Int J Mol Med 47:N/A (2021). PubMed: 33693953
    • Yenerall P  et al. RUVBL1/RUVBL2 ATPase Activity Drives PAQosome Maturation, DNA Replication and Radioresistance in Lung Cancer. Cell Chem Biol 27:105-121.e14 (2020). PubMed: 31883965

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