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Anti-ATR抗体(ab10312)

Submit a review Q&A (2)References (7)
Western blot - Anti-ATR antibody (ab10312)
  • Western blot - Anti-ATR antibody (ab10312)

Key features and details

  • Rabbit polyclonal to ATR
  • Suitable for: WB
  • Reacts with: Human
  • Isotype: IgG

概述

  • 产品名称

    Anti-ATR抗体
    参阅全部 ATR 一抗

  • 描述

    兔多克隆抗体to ATR

  • 宿主

    Rabbit

  • 特异性

    We have data to indicate that this antibody may not cross react with Xenopus laevis. However, this has not been conclusively tested and expression levels may vary in certain cell lines/tissues.

  • 经测试应用

    适用于: WBmore details

  • 种属反应性

    与反应: Human
    预测可用于: Chimpanzee, Rhesus monkey, Gorilla, Orangutan

  • 免疫原

    Synthetic peptide within Human ATR aa 400-500. The exact immunogen sequence used to generate this antibody is proprietary information. If additional detail on the immunogen is needed to determine the suitability of the antibody for your needs, please contact our Scientific Support team to discuss your requirements.
    Database link: Q13535

  • 阳性对照

    • Whole cell lysate from HeLa and U2OS cells

  • 常规说明

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. 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, along with publications, customer reviews and Q&As

性能

应用

The Abpromise guarantee

Abpromise™承诺保证使用ab10312于以下的经测试应用

“应用说明”部分 下显示的仅为推荐的起始稀释度；实际最佳的稀释度/浓度应由使用者检定。

应用 Ab评论 说明
WB
1/15000 - 1/30000. Detects a band of approximately 300 kDa (predicted molecular weight: 317 kDa).
说明
WB
1/15000 - 1/30000. Detects a band of approximately 300 kDa (predicted molecular weight: 317 kDa).

靶标

  • 功能

    Serine/threonine protein kinase which activates checkpoint signaling upon genotoxic stresses such as ionizing radiation (IR), ultraviolet light (UV), or DNA replication stalling, thereby acting as a DNA damage sensor. Recognizes the substrate consensus sequence [ST]-Q. Phosphorylates BRCA1, CHEK1, MCM2, RAD17, RPA2, SMC1 and p53/TP53, which collectively inhibit DNA replication and mitosis and promote DNA repair, recombination and apoptosis. Phosphorylates 'Ser-139' of histone variant H2AX/H2AFX at sites of DNA damage, thereby regulating DNA damage response mechanism. Required for FANCD2 ubiquitination. Critical for maintenance of fragile site stability and efficient regulation of centrosome duplication.

  • 组织特异性

    Ubiquitous, with highest expression in testis. Isoform 2 is found in pancreas, placenta and liver but not in heart, testis and ovary.

  • 疾病相关

    Defects in ATR are a cause of Seckel syndrome type 1 (SCKL1) [MIM:210600]. SCKL1 is a rare autosomal recessive disorder characterized by growth retardation, microcephaly with mental retardation, and a characteristic 'bird-headed' facial appearance.

  • 序列相似性

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

  • 翻译后修饰

    Phosphorylated; autophosphorylates in vitro.

  • 细胞定位

    Nucleus. Nucleus > PML body. Depending on the cell type, it can also be found in PML nuclear bodies. Recruited to chromatin during S-phase. Redistributes to discrete nuclear foci upon DNA damage, hypoxia or replication fork stalling.
  • Information by UniProt

  • 数据库链接

    • 别名

      • Ataxia telangiectasia and Rad3 related antibody
      • Ataxia telangiectasia and Rad3-related protein antibody
      • ATR antibody
      • ATR_HUMAN antibody
      • FCTCS antibody
      • FRAP Related Protein 1 antibody
      • FRAP-related protein 1 antibody
      • FRP1 antibody
      • MEC1 antibody
      • MEC1 mitosis entry checkpoint 1 homolog antibody
      • Protein kinase ATR antibody
      • RAC3 antibody
      • Rad3 related protein antibody
      • SCKL antibody
      • SCKL1 antibody
      • Serine/threonine protein kinase ATR antibody
      • Serine/threonine-protein kinase ATR antibody
      see all

    图片

    • Western blot - Anti-ATR antibody (ab10312)
      Western blot - Anti-ATR antibody (ab10312)
      All lanes : Anti-ATR antibody (ab10312) at 1/15000 dilution

      Lane 1 : Wild-type A549 cell lysate
      Lane 2 : ATR knockout A549 cell lysate
      Lane 3 : HeLa cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 317 kDa
      Observed band size: 260 kDa why is the actual band size different from the predicted?



      False colour image of Western blot: Anti-ATR antibody staining at 1/15000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab10312 was shown to bind specifically to ATR. A band was observed at 260 kDa in wild-type A549 cell lysates with no signal observed at this size in ATR knockout cell line ab276104 (knockout cell lysate ab277987). To generate this image, wild-type and ATR 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 3 % 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-ATR antibody (ab10312)
      Western blot - Anti-ATR antibody (ab10312)
      Detection of human ATR by Western Blot. Samples: Whole cell lysate (20 µg) from HeLa and U2OS cells separated on a 3 to 8% tris-acetate gel. Antibody: ab10312 used at 0.07 mg/ml. Detection: Chemiluminescence with 15 second exposure. Detection of human ATR by Western Blot. Samples: Whole cell lysate (20 µg) from HeLa and U2OS cells separated on a 3 to 8% tris-acetate gel. Antibody: ab10312 used at 0.07 mg/ml. Detection: Chemiluminescence with 15 second exposure.

    文献 (7)

    发表研究结果有使用 ab10312？请让我们知道，以便我们可以引用本数据表中的参考文章。

    ab10312 被引用在 7 文献中.

    • Bianco JN  et al. Overexpression of Claspin and Timeless protects cancer cells from replication stress in a checkpoint-independent manner. Nat Commun 10:910 (2019). PubMed: 30796221
    • Xia Y  et al. Garcinone C exerts antitumor activity by modulating the expression of ATR/Stat3/4E-BP1 in nasopharyngeal carcinoma cells. Oncol Rep 39:1485-1493 (2018). PubMed: 29344638
    • Zhang P  et al. ATM-mediated stabilization of ZEB1 promotes DNA damage response and radioresistance through CHK1. Nat Cell Biol 16:864-75 (2014). PubMed: 25086746
    • Chen J  et al. Repair of naturally occurring mismatches can induce mutations in flanking DNA. Elife 3:e02001 (2014). PubMed: 24843013
    • Sousa MM  et al. An inverse switch in DNA base excision and strand break repair contributes to melphalan resistance in multiple myeloma cells. PLoS One 8:e55493 (2013). WB ; Human . PubMed: 23405159
    • Treanor L  et al. Independent regulation of P53 stabilisation and activation after Rb deletion in primary epithelial cells. Int J Oncol 37:31-9 (2010). ICC/IF, WB ; Mouse . PubMed: 20514394
    • Derheimer FA  et al. RPA and ATR link transcriptional stress to p53. Proc Natl Acad Sci U S A 104:12778-83 (2007). PubMed: 17616578

    客户评价及客户问答

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    1-2 of 2 Abreviews or Q&A

    Question

    BATCH NUMBER 160963 ORDER NUMBER -- NOT SPECIFIED -- DESCRIPTION OF THE PROBLEM too many bands and not any specific SAMPLE whole cell extracts (human glioblastoma cell lines) PRIMARY ANTIBODY Abcam antiATR, rabbit, ab10312, lot160963, dilution 1/15000 in PBS, BSA 1%, 2hr Room Temp, several 15 min washes in PBS-tween DETECTION METHOD ECL POSITIVE AND NEGATIVE CONTROLS USED very good loading control (actin) so it is nt an extract or membrane or transfer or detection problem, but truely a problem of primiray Ab (long experience of western in the lab, main technic used) ANTIBODY STORAGE CONDITIONS 4°C SAMPLE PREPARATION standard (protease inhibitor cocktail-Roche, heated in 1X loading buffer (SDS, DTT, blue) AMOUNT OF PROTEIN LOADED 60 µg ELECTROPHORESIS/GEL CONDITIONS reducing, 10% TRANSFER AND BLOCKING CONDITIONS ON transfer, TG iX, 15 % Ethanol, PVDF membrane, blocking 5% non fat milk SECONDARY ANTIBODY antirabbit-HRP (jackson) 1/5000 in PBS-T, 1hr RT, several 15 min washes in PBS-tween HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 3 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? increase the dilution because background whorse at 1/5000 and 1/10000

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    Answer

    Thank you for taking the time to fill out the technical questionaire and send it in to us. I apologise for the delay in getting back to you. However, I have more details of the western blot testing procedure for this antibody from the laboratory which I hope will help. This antibody can often recognise extra bands which are fragments of ATR, although this has yet to be tested on glioblastoma cells. If the small bands are more prevalent relative to the full-size ATR in this cell line, there may be a problem which we can resolve by optimisation. 1. The small bands may represent fragments of ATR. Ensure the lysate is fresh and correctly prepared. Ensure samples are adequately denatured and reduced. 2. The gels should be of lower percentage, 4 to 8%. 3. The primary antibody is at a high concentration which may lead to non specific binding. The data sheet suggests using a dilution of 1:15000 to 1:30000. There has been an Abreview submitted for this antibod in which the antibody was diluted 1:50000 with good results. 4. The amount of protein loaded is quite high. Try 20-30ug per lane. 5. You can try increasing the blocking incubation time to 2-3 hours. When the antibody was tested the laboratory, they used used 5% Non-fat Dry Milk in TBS with 0.05% Tween to block the membrane. The same was used as diluent for the primary and secondary antibody. I hope this is helpful information and that these suggestions will improve the results. Should you still obtain an unsatisfactory result after using these suggestions, then please do not hesitate to get back to me.

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    Question

    my Mphile project is to concentrate on an ATM/ATR DNA damage model, and i am thinking to order the ATM antibody [2B5] (ab4471) and ATR antibody (ab10312) that used in flow cytometry, western blot and immunohistochemistry from your company.in order to make sure the satisfy of my result the quality of these antibodies are very important, so there are few questions i would like to ask and would be greatful if you could reply me as well. first, is that your ATR [2B5] antibody cross reacts with ATM (ATX08) antibody? second, same as above is that your ATM cross reacts with ATR? it is really important that there is no cross reaction between these two antibodies as i want to exam ATM and ATR separately. third. does your ATM [2B5] antibody works on western blot? forth, the western blot diagram on the datasheet of ATR [2b5], could u explain to us in details about this diagram please, as there are two bands on this diagram, is that means your ATM and ATR antibody are corss react with each other, if so we could not order it as we do not want the ATM and ATR antibody cross react. please e mail us back as soon as possible. look forward to hearing from u

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    Answer

    You recently enquired about our ATM and ATR antibodies and my colleague Karen has already sent you a reply regarding the antibodies ab2354 and ab4471. I have also investigated your enquiry regarding ab10312. The antibody should not react with ATM owing to complete divergence of amino acid sequence between ATM and ATR at the site to which the epitope maps. There is no indication that the antibody crossreacts with ATM. I hope this further information helps,

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