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遺伝子ノックアウト細胞株 検証済リコンビナントRabMAb

Anti-p95/NBS1 抗体 [Y112] (ab32074)

Reviews (5)Q&A (7)References (26)
Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
  • Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
  • Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p95/NBS1 antibody [Y112] (ab32074)
  • Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p95/NBS1 antibody [Y112] (ab32074)
  • Anti-p95/NBS1 antibody [Y112] (ab32074)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [Y112] to p95/NBS1
  • Suitable for: WB, IHC-P
  • Knockout validated
  • Reacts with: Human

Conjugates logo Related conjugates and formulations

Carrier Free

製品の概要

  • 製品名

    Anti-p95/NBS1 antibody [Y112]
    p95/NBS1 一次抗体 製品一覧

  • 製品の詳細

    Rabbit monoclonal [Y112] to p95/NBS1

  • 由来種

    Rabbit

  • アプリケーション

    適用あり: WB, IHC-Pmore details
    適用なし: Flow Cyt or ICC/IF

  • 種交差性

    交差種: Human
    交差が予測される動物種: Mouse, Rat

  • 免疫原

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • ポジティブ･コントロール

    • WB: HeLa, Jurkat and Wild-type A431 cell lysates. IHC-P: Human testis and skin carcinoma tissues.

  • 特記事項

    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.

製品の特性

アプリケーション

The Abpromise guarantee

Abpromise保証は、 次のテスト済みアプリケーションにおけるab32074の使用に適用されます

アプリケーションノートには、推奨の開始希釈率がありますが、適切な希釈率につきましてはご検討ください。

アプリケーション Abreviews 特記事項
WB (3)
1/1000 - 1/10000. Predicted molecular weight: 85 kDa.

For unpurified use at 1/1000 - 1/2000.
IHC-P (1)
1/200. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

For unpurified use at 1/50.
特記事項
WB
1/1000 - 1/10000. Predicted molecular weight: 85 kDa.

For unpurified use at 1/1000 - 1/2000.
IHC-P
1/200. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

For unpurified use at 1/50.
追加情報
Is unsuitable for Flow Cyt or ICC/IF.

ターゲット情報

  • 機能

    Component of the MRE11-RAD50-NBN (MRN complex) which plays a critical role in the cellular response to DNA damage and the maintenance of chromosome integrity. The complex is involved in double-strand break (DSB) repair, DNA recombination, maintenance of telomere integrity, cell cycle checkpoint control and meiosis. The complex possesses single-strand endonuclease activity and double-strand-specific 3'-5' exonuclease activity, which are provided by MRE11A. RAD50 may be required to bind DNA ends and hold them in close proximity. NBN modulate the DNA damage signal sensing by recruiting PI3/PI4-kinase family members ATM, ATR, and probably DNA-PKcs to the DNA damage sites and activating their functions. It can also recruit MRE11 and RAD50 to the proximity of DSBs by an interaction with the histone H2AX. NBN also functions in telomere length maintenance by generating the 3' overhang which serves as a primer for telomerase dependent telomere elongation. NBN is a major player in the control of intra-S-phase checkpoint and there is some evidence that NBN is involved in G1 and G2 checkpoints. The roles of NBS1/MRN encompass DNA damage sensor, signal transducer, and effector, which enable cells to maintain DNA integrity and genomic stability. Forms a complex with RBBP8 to link DNA double-strand break sensing to resection. Enhances AKT1 phosphorylation possibly by association with the mTORC2 complex.

  • 組織特異性

    Ubiquitous. Expressed at high levels in testis.

  • 関連疾患

    Nijmegen breakage syndrome
    Breast cancer
    Aplastic anemia
    Defects in NBN might play a role in the pathogenesis of childhood acute lymphoblastic leukemia (ALL).

  • 配列類似性

    Contains 1 BRCT domain.
    Contains 1 FHA domain.

  • ドメイン

    The FHA and BRCT domains are likely to have a crucial role for both binding to histone H2AFX and for relocalization of MRE11/RAD50 complex to the vicinity of DNA damage.
    The C-terminal domain contains a MRE11-binding site, and this interaction is required for the nuclear localization of the MRN complex.
    The EEXXXDDL motif at the C-terminus is required for the interaction with ATM and its recruitment to sites of DNA damage and promote the phosphorylation of ATM substrates, leading to the events of DNA damage response.

  • 翻訳後修飾

    Phosphorylated by ATM in response of ionizing radiation, and such phosphorylation is responsible intra-S phase checkpoint control and telomere maintenance.

  • 細胞内局在

    Nucleus. Nucleus, PML body. Chromosome, telomere. Localizes to discrete nuclear foci after treatment with genotoxic agents.
  • Information by UniProt

  • 参照データベース

    see all

  • 別名

    • AT V1 antibody
    • AT V2 antibody
    • ATV antibody
    • Cell cycle regulatory protein p95 antibody
    • FLJ10155 antibody
    • MGC87362 antibody
    • Nbn antibody
    • NBN_HUMAN antibody
    • NBS 1 antibody
    • NBS antibody
    • NBS1 antibody
    • Nibrin antibody
    • Nijmegen breakage syndrome 1 (nibrin) antibody
    • Nijmegen breakage syndrome antibody
    • Nijmegen breakage syndrome protein 1 antibody
    • p95 antibody
    • p95 protein of the MRE11/RAD50 complex antibody
    see all

画像

  • Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
    Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
    All lanes : Anti-p95/NBS1 antibody [Y112] (ab32074) at 1/1000 dilution

    Lane 1 : Wild-type A431 cell lysate
    Lane 2 : NBN knockout A431 cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 85 kDa
    Observed band size: 90 kDa why is the actual band size different from the predicted?



    Lanes 1 - 2: Merged signal (red and green). Green - ab32074 observed at 90 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.

    ab32074 was shown to react with p95/NBS1 in wild-type A431 cells in Western blot with loss of signal observed in NBN knockout cell line ab269506 (NBN knockout cell lysate ab269668). Wild-type A431 and NBN knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5 % milk in TBS-T (0.1 % Tween®) before incubation with ab32074 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 h at room temperature before imaging.

  • Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
    Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
    All lanes : Anti-p95/NBS1 antibody [Y112] (ab32074) at 1/1000 dilution

    Lane 1 : Wild-type HeLa cell lysate
    Lane 2 : NBN knockout HeLa cell lysate
    Lane 3 : Wild-type HeLa nuclear cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 85 kDa
    Observed band size: 95 kDa why is the actual band size different from the predicted?



    Lanes 1-3: Merged signal (red and green). Green - ab32074 observed at 95 kDa.

    ab32074 Anti-p95/NBS1 antibody [Y112] was shown to specifically react with p95/NBS1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab261834 (knockout cell lysate ab257111) was used. Wild-type and p95/NBS1 knockout samples were subjected to SDS-PAGE. ab32074, Anti-GAPDH antibody [6C5] - Cytoplasmic Loading Control (ab8245) and Anti-Histone H3 (ab176842) - Nuclear Loading Control were incubated overnight at 4°C at 1 in 1000 dilution, 1 in 20000 dilution and 1 in 1000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773), Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) at 1 in 10000 dilution for 1 hour at room temperature before imaging.

     

     

  • Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
    Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
    All lanes : Anti-p95/NBS1 antibody [Y112] (ab32074) at 1/1200 dilution (unpurified)

    Lane 1 : HeLa cell lysate
    Lane 2 : Jurkat cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Peroxidase-conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

    Predicted band size: 85 kDa
    Observed band size: 95 kDa why is the actual band size different from the predicted?



    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p95/NBS1 antibody [Y112] (ab32074)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p95/NBS1 antibody [Y112] (ab32074)

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human testis tissue labelling p95/NBS1 with purified ab32074 at 1/200. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. A prediluted HRP-polymer conjugated anti-rabbit IgG was used as the secondary antibody. Counterstained with hematoxylin.

  • Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
    Western blot - Anti-p95/NBS1 antibody [Y112] (ab32074)
    All lanes : Anti-p95/NBS1 antibody [Y112] (ab32074) at 1/10000 dilution (purified)

    Lane 1 : HeLa cell lysate
    Lane 2 : Jurkat cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Peroxidase-conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

    Predicted band size: 85 kDa
    Observed band size: 95 kDa why is the actual band size different from the predicted?



    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p95/NBS1 antibody [Y112] (ab32074)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p95/NBS1 antibody [Y112] (ab32074)

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human testis tissue labelling p95/NBS1 with unpurified ab32074 at 1/20. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. A prediluted HRP-polymer conjugated anti-rabbit IgG was used as the secondary antibody. Counterstained with hematoxylin.

  • Anti-p95/NBS1 antibody [Y112] (ab32074)
    Anti-p95/NBS1 antibody [Y112] (ab32074)

プロトコール

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

参考文献 (26)

ab32074 を使用した論文を発表された方は、こちらまでお知らせください。データシートに掲載させていただきます。

ab32074 は 26 報の論文で使用されています。

  • Zhang B  et al. NBS1 is required for SPO11-linked DNA double-strand break repair in male meiosis. Cell Death Differ 27:2176-2190 (2020). PubMed: 31965061
  • Dellino GI  et al. Release of paused RNA polymerase II at specific loci favors DNA double-strand-break formation and promotes cancer translocations. Nat Genet 51:1011-1023 (2019). PubMed: 31110352
  • Leimbacher PA  et al. MDC1 Interacts with TOPBP1 to Maintain Chromosomal Stability during Mitosis. Mol Cell 74:571-583.e8 (2019). PubMed: 30898438
  • Carvajal-Maldonado D  et al. Perturbing cohesin dynamics drives MRE11 nuclease-dependent replication fork slowing. Nucleic Acids Res 47:1294-1310 (2019). PubMed: 29917110
  • Afzal S  et al. Rat Glioma Cell-Based Functional Characterization of Anti-Stress and Protein Deaggregation Activities in the Marine Carotenoids, Astaxanthin and Fucoxanthin. Mar Drugs 17:N/A (2019). PubMed: 30909572
View all Publications for this product

レビューと Q&A

Show All レビュー Q&A
レビューを送る 質問を送る

1-10 of 12 Abreviews or Q&A

Western blot abreview for Anti-p95 NBS1 antibody [Y112]

 Excellent
Abreviews
abreview image
Application
Western blot
Sample
Human Cell lysate - whole cell (HeLa)
Gel Running Conditions
Reduced Denaturing (4-12% Bis-Tris)
Loading amount
50 µg
Treatment
10 Gy IR
Specification
HeLa
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

投稿 Apr 24 2013

Immunocytochemistry/ Immunofluorescence abreview for Anti-p95 NBS1 antibody [Y112]

 Excellent
Abreviews
This product is known to not work in this application or species.
abreview image
Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HeLa)
Permeabilization
Yes - Triton X-100
Specification
HeLa
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5%
Fixative
Formaldehyde
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

投稿 May 30 2014

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-p95/NBS1 antibody [Y112]

 Excellent
Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Human carotid plaque tissue)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citrate buffer
Permeabilization
No
Specification
Human carotid plaque tissue
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative
Formaldehyde
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

投稿 Nov 23 2011

Western blot abreview for Anti-p95 NBS1 antibody [Y112]

 Excellent
Abreviews
abreview image
Application
Western blot
Sample
Human Cell lysate - whole cell (Jurkat cells)
Gel Running Conditions
Reduced Denaturing
Loading amount
100000 cells
Specification
Jurkat cells
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

投稿 May 19 2010

Question

Inquiry: i wanted to know the concentration of tha antibody Anti-p95 NBS1 ChIP grade (ab32074) lot: GR 4228-7 thanks in advance

Read More
Answer

Thank you for your inquiry.

I am happy to confirm that the concentration of lot GR4228-7 is 0.645 mg/mL.

I also would like to comment on the seemingly low concentration:

The IgG produced by rabbits have greater affinity and greater sensitivity than mouse IgG (about 100-1000 fold) and at the same time the total amount of IgG produced by the rabbit hybridomas is generally lower than in mouse hybridomas.

Many 'veteran' antibody users are generally familiar with the IgG amount provided from mouse antibodies and can judge how many reactions they may get or need to use based on concentration. Unfortunately, this experience is not transferable to rabbit monoclonal antibodies because the concentration scales are generally very different.

Since the specificity and sensitivity is much higher with rabbit monoclonal antibodies, less IgG is required per reaction.

I would like to reassure you that while the amount of IgG may not be comparable to mouse monoclonal antibodies, customers are receiving a comparable number of reactions/tests/blots/stainings per tube.

I hope this information is helpful. Please do not hesitate to contact me with any further questions in this matter.

Read More

Question

what is the concentration of Anti-p95 NBS1 antibody [Y112] - ChIP Grade (ab32074) lot GR4228-5?

Read More
Answer


The concentration of ab32074 lot GR4228-5is : 0.645 mg/mL.

Read More

Question

Batch variability in ChIP. Would like to replace vials from alot that did not work well in ChIPwith vials from adifferent lot.

Read More
Answer

Thank you for your call today and for keeping me updated about this situation.

I am sending the three vials thatwe havein stock. These vials will be arriving on the free of charge order 1060035, and I expect them to arrive tomorrow.

I've also issued credit note***for one vial of ab32074. The full purchase price will be refunded back to your credit card pending approval from our accounting department. If you have any questions about this credit note, please contact mailto:us.credits@abcam.com.

I apologize again for the batch variability of this antibody. Please let me know if you have any questions or if there is anything else that we can do for you.

Read More

Question

Dear Madam / Sir,

A user needs a information about positive control tissue p95 NBS1 antibody [Y112] (ab32074). It is stated that Human skin carcinoma as positive control tissue on the datasheet. But It isn't clear which types of skin cancer. Could you please clear that?

Thank you.

Best regards,

Read More
Answer

Thank you for your enquiry.

I can confirm that p95 NBS1 antibody [Y112] (ab32074) was tested onsquamous cell carcinoma of the skin. This would be a suitable positive control.

I hope this information will be helpful to you. If you have any further questions, please do not hesitate to contact us.

Read More

Question

What is the concentration of this antibody, lot #GR50908-2?

Read More
Answer

Thank you for contacting Abcam.



The concentration for ab32074, lot #GR50908-2 is *****mg/mL.



Please let me know if there is anything else I can help you with.

Read More

Question

Lot GR4228-7 showed enrichment for negative control gene with mouse lymphocyte samples. Is lot GR50908-2 better for ChIP?

Read More
Answer

Thanks for your call today and for letting us know about the trouble with ab32074 in ChIP.

As we discussed, I am sending a free of charge vial of ab32074 from lot GR50908-2 on the order ***, which should arrive tomorrow.

I've looked through our records, and we do not test this antibody in-house in ChIP. This application was added to our datasheet based on results in the following article-

http://www.ncbi.nlm.nih.gov/pubmed/21255732?dopt=Abstract

So unfortunately I don't have any data for each of these lots in ChIP, but I am hopeful that the new vial will work better for you. Please keep me updated about the results with this vial, and let me know if you have any questions or need anything else from me.

Read More

1-10 of 12 Abreviews or Q&A

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