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  1. Link

    metnase-antibody-ab3823.pdf

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Epigenetics and Nuclear Signaling Chromatin Modifying Enzymes Methylation
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Anti-Metnase 抗体 (ab3823)

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Submit a review Q&A (3)References (4)

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Western blot - Anti-Metnase antibody (ab3823)

    Key features and details

    • Goat polyclonal to Metnase
    • Suitable for: WB
    • Reacts with: Human
    • Isotype: IgG

    こちらの製品もご検討ください

    タンパク質
    Product image
    Recombinant Human Metnase protein (ab125543)
    一次抗体
    Product image
    Anti-ZDHHC13 antibody (ab28759)
    二次抗体
    Product image
    Donkey Anti-Goat IgG H&L (HRP) (ab97110)

    関連製品

    製品の概要

    • 製品名

      Anti-Metnase antibody
      Metnase 一次抗体 製品一覧
    • 製品の詳細

      Goat polyclonal to Metnase
    • 由来種

      Goat
    • アプリケーション

      適用あり: WBmore details
    • 種交差性

      交差種: Human
    • 免疫原

      Synthetic peptide corresponding to Human Metnase aa 650 to the C-terminus (C terminal).
      Database link: NP_006506.3

      Run BLAST with BLAST the sequence with ExPASy Run BLAST with BLAST the sequence with NCBI
    • ポジティブ・コントロール

      • Daudi lysate.
    • 特記事項

      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

    製品の特性

    • 製品の状態

      Liquid
    • 保存方法

      Shipped at 4°C. Upon delivery aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
    • バッファー

      pH: 7.30
      Preservative: 0.02% Sodium azide
      Constituents: 0.5% Tris, 0.5% BSA
    • Concentration information loading...
    • 精製度

      Immunogen affinity purified
    • 特記事項(精製)

      Purified from goat serum by ammonium sulphate precipitation followed by antigen affinity chromatography using the immunizing peptide.
    • ポリ/モノ

      ポリクローナル
    • アイソタイプ

      IgG
    • 研究分野

      • Epigenetics and Nuclear Signaling
      • Chromatin Modifying Enzymes
      • Methylation
      • Epigenetics and Nuclear Signaling
      • Chromatin Modifying Enzymes
      • Methylation
      • Lysine methylation

    関連製品

    • Compatible Secondaries

      • Donkey Anti-Goat IgG H&L (Alexa Fluor® 488) (ab150129)
      • Donkey Anti-Goat IgG H&L (HRP) (ab205723)
    • Isotype control

      • Goat IgG, polyclonal - Isotype Control (ab37373)
    • Recombinant Protein

      • Recombinant Human Metnase protein (ab125543)

    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ab3823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
    Use a concentration of 1 - 3 µg/ml. Detects a band of approximately 70-75 kDa.

    A 1 hour primary incubation is recommended for this product. Approx 70 - 75 kDa band seen in Daudi lysate.

    特記事項
    WB
    Use a concentration of 1 - 3 µg/ml. Detects a band of approximately 70-75 kDa.

    A 1 hour primary incubation is recommended for this product. Approx 70 - 75 kDa band seen in Daudi lysate.

    ターゲット情報

    • 機能

      Histone methyltransferase that methylates 'Lys-4' and 'Lys-36' of histone H3, 2 specific tags for epigenetic transcriptional activation. Specifically mediates dimethylation of H3 'Lys-36'. Has sequence-specific DNA-binding activity and recognizes the 19-mer core of the 5'-terminal inverted repeats (TIRs) of the Hsmar1 element. Has DNA nicking activity. Has in vivo end joining activity and may mediate genomic integration of foreign DNA.
    • 組織特異性

      Widely expressed, with highest expression in placenta and ovary and lowest expression in skeletal muscle.
    • 配列類似性

      In the N-terminal section; belongs to the histone-lysine methyltransferase family.
      In the C-terminal section; belongs to the mariner transposase family.
      Contains 1 post-SET domain.
      Contains 1 pre-SET domain.
      Contains 1 SET domain.
    • ドメイン

      The mariner transposase Hsmar1 region mediates DNA-binding. It has no transposase activity because the active site contains an Asn in position 610 instead of a Asp residue.
    • 細胞内局在

      Nucleus. Chromosome.
    • Target information above from: UniProt accession Q53H47 The UniProt Consortium
      The Universal Protein Resource (UniProt) in 2010
      Nucleic Acids Res. 38:D142-D148 (2010) .

      Information by UniProt
    • 参照データベース

      • Entrez Gene: 6419 Human
      • Omim: 609834 Human
      • SwissProt: Q53H47 Human
      • Unigene: 716054 Human
      • 別名

        • Histone lysine N methyltransferase antibody
        • Histone lysine N methyltransferase SETMAR antibody
        • Hsmar 1 antibody
        • Hsmar1 antibody
        • Mariner transposase Hsmar1 antibody
        • Metnase antibody
        • SET domain and mariner transposase fusion antibody
        • SET domain and mariner transposase fusion gene antibody
        • SET domain and mariner transposase fusion gene containing protein antibody
        • SET domain and mariner transposase fusion gene-containing protein antibody
        • Setmar antibody
        • SETMR_HUMAN antibody
        see all

      画像

      • Western blot - Anti-Metnase antibody (ab3823)
        Western blot - Anti-Metnase antibody (ab3823)
        ab3823 staining (2µg/ml) of Daudi lysate (RIPA buffer, 30µg total protein per lane).  Primary incubated for 12 hours.  Detected by western blot using chemiluminescence. ab3823 staining (2µg/ml) of Daudi lysate (RIPA buffer, 30µg total protein per lane). Primary incubated for 12 hours. Detected by western blot using chemiluminescence.

      プロトコール

      • Western blot protocols

      Click here to view the general protocols

      データシートおよび資料

      • Datasheet download

        Download

      参考文献 (4)

      ab3823 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。

      ab3823 は 4 報の論文で使用されています。

      • Tellier M & Chalmers R Compensating for over-production inhibition of the Hsmar1 transposon in Escherichia coli using a series of constitutive promoters. Mob DNA 11:5 (2020). PubMed: 31938044
      • Sharma N  et al. Distinct roles of structure-specific endonucleases EEPD1 and Metnase in replication stress responses. NAR Cancer 2:zcaa008 (2020). PubMed: 32743552
      • Tellier M & Chalmers R The roles of the human SETMAR (Metnase) protein in illegitimate DNA recombination and non-homologous end joining repair. DNA Repair (Amst) 80:26-35 (2019). PubMed: 31238295
      • Van Duyne R  et al. Lysine methylation of HIV-1 Tat regulates transcriptional activity of the viral LTR. Retrovirology 5:40 (2008). WB ; Human . PubMed: 18498648

      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

      1-3 of 3 Abreviews or Q&A

      Question

      Here is the information you requested. Lot #15383, Invoice # 12808 > 1. Please describe the problem (high background, wrong band size, more > bands, no band etc). In all Westerns using the primary antibody, multiple bands are present; in fact, when compared to a Coomassie stained gel, all bands visible in the Coomassie gel also appear to be stained in the Western blot. > > 2. On what material are you testing the antibody in WB? >Species? Human > Cell extract/ Nuclear extract? We have tested two independent cell extracts: an Imgenex Daudi lysate and an ATCC Daudi cell line we maintain in culture > Purified protein? No, crude extract > Recombinant protein? No > 3. How much protein did you load? 30 micrograms (according to Abcam protocol) > How did you prepare the lysate for the analysis (protease inhibitors etc)? The Imgenex Daudi cell lysate is ready-to-load, it comes with beta-mercaptoethanol and sample buffer added. The Daudi cell culture we maintain was prepared according to the Abcam protocol we received. >Did you heat the samples? No, not necessary for Imgenex cell lysate. Yes, for the Daudi cell culture we maintain (according to Abcam protocol) > > 4. Primary Antibody > Specification (in which species was it raised against)? Goat > At what dilution(s) have you tested this antibody? 0.5ug/ml, 1.0ug/ml, and 2ug/ml (2ug/ml is recommended in Abcam protocol) > Incubation time, wash step? Tested incubation time at both 1hr and overnight. Wash in 1X PBST (Tween tested at both 0.1% and 0.05%) 3X for 5-15min > > 5. Secondary Antibody > Specification (in which species was it raised against)? Rabbit anti-goat IgG purchased from Sigma (A-8919) > At what dilution(s) have you tested this antibody? 1:3000 and 1:4000 > Incubation time, wash step? 1hr, same washes tested as for primary antibody > Do you know whether the problems you are experiencing come from the > secondary? We have performed a "no primary" control, and there is almost no staining present. > > 6. What detection method are you using? HRP with DAB, metal-enhanced, developed 5-10 min. > > 7. Background bands > Have you eliminated the possibility that any background bands could > be due to the secondary antibody? (Run a â??No primaryâ?? control) Yes Is the blocking step sufficient? (We recommend blocking the membrane by > adding 20 ml of blocking buffer (5% non-fat dry milk, 0.1% Tween-20 in > TBS). Incubate for 2 h at room temperature or overnight at 4°C with > agitation) We use 5% non-fat dry milk, 0.1% Tween-20 in PBS overnight at 4C. Are your washing steps sufficiently stringent? (Multiple > short washes are more effective than fewer longer wash steps) We have washed 3X for 15 minutes each. >At what size are the bands migrating? Could they be degradation products of your target? It appears that all bands in the lysate are stained in the Western. >Please provide an image of your blot (as an e-mail attachment, a faxed image is not sufficient) The attached image shows 30ug and 20ug of Imgenex Daudi lysate using the SETMAR primary antibody. The far left lane is our ladder, which doesn't hold up too well during the processing, but we marked the approximate positions of 39kD and 78kD. The SETMAR primary antibody should identify a 75kD protein. The far right lane is our positive control which is described in detail below. > 8. Optimization attempts How many times have you tried the Western? Seven. Do you obtain the same results every time e.g. are background bands > always in the same place? Yes What steps have you altered? We have altered primary antibody concentrations, secondary antibody concentrations, incubation times, number of washes and length of time, Tween concentration, and lysate mass loaded. > > 9. Did you apply positive and negative controls along with the samples? > Please specify. Our positive control is alpha-actinin purified protein from chicken gizzard (Sigma A9776) detected with monoclonal anti-alpha-actinin from mouse (Sigma A5044), using an anti-mouse secondary antibody (Sigma) A9917.

      Read More

      Abcam community

      Verified customer

      Asked on May 10 2004

      Answer

      Thank you very much for your patience. The only suggestion that I have at this point (you have done pretty much everything!) is to try blocking with 3% BSA instead. The originator didn't see high background with this antibody and used a more sensitive detection system (ECL plus). Did you buy the antibody directly from us or from a distributor, such as Novus Biologicals? And what was the date? I'm not coming up with a record of this antibody being bought from your school. Thanks...

      Read More

      Abcam Scientific Support

      Answered on May 24 2004

      Question

      Customer's question: I am trying to replicate the Western blot results for ab3823 SETMAR antibody shown on the Abcame website. I received a copy of the protocol and I still have some questions. Could you tell me the boiling temperature at which you incubated the RIPA buffer lysate plus the 2x SDS sample buffer? Thanks for your help!

      Read More

      Abcam community

      Verified customer

      Asked on Nov 20 2003

      Answer

      The samples (in 1.5ml microtubes) were simply put into boiling water. So I would expect the temperature to be almost exactly 100 degrees centigrade.

      Read More

      Abcam Scientific Support

      Answered on Nov 21 2003

      Question

      Dear tech support, We are trying to replicate the Western blot results for ab3823 SETMAR antibody shown on the Abcam website. Could you please send us a detailed protocol for your Western blots?

      Read More

      Abcam community

      Verified customer

      Asked on Oct 02 2003

      Answer

      As stated on the datasheet we used the following conditions to get our result: ab3823 staining (2µg/ml) of Daudi lysate (RIPA buffer, 30µg total protein per lane). Primary incubated for 12 hours. Detected by western blot using chemiluminescence However additional information about our protocol is given below: - Lysis. Cell pellets were washed with ice-cold PBS. 1 ml of RIPA buffer was added per 1E8 cells and incubated on ice for 20 min, vortexing 2-3 times, briefly. The lysate was aliquotted into 1.5 ml microfuge tubes and centrifuged at 13,000 rpm for 5 min in a microfuge. The supernatant was transferred into clean tubes and its protein concentration was measured with BioRad protein assay. The concentration was then adjusted to 5 mg/ml with RIPA lysis buffer. An equal volume of 2 x SDS sample buffer was then added and the cell lysate was boiled for 5 minutes. Lysates were stored at -80C until use.(RIPA buffer = 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1 mM PMSF, 1 mM EDTA, 5 µg/ml Aprotinin, 5 µg/ml Leupeptin, 1% Triton -100, 1% Sodium deoxycholate, 0.1% SDS). - SDS PAGE. Samples were run at 200V constant on a 12% acrylamide SDS-PAGE mini gel - using Biorad Mini-Protean 3 kit and protocols. Before loading samples had 5% (v/v) 2-ME added and were boiled for 3 minutes. - Transfer. We used a Biorad Mini Trans-Blot, constant 100 V for 1 hour. Transfer Buffer was 20 mM Tris pH 8.0, 150 mM Glycine, 10% Methanol. We transferred to Millipore PVDF membrane and stained with Ponceau Red to evaluate the transfer. - Staining. The membrane was blocked in 2.5% skimmed milk in TBS-T (TBS + 0.05% Tween-20) for 1 hr at room temperature with agitation. Primary antibody was incubated for 1 hr at room temperature with agitation. We used anti-goat-HRP at 1:3000 for 1 hr at room temperature with agitation. We washed with TBST three times after primary and secondary antibody, each wash lasting for 5-10 mins. ECL-plus (Amersham) was used rather than ECL, which is considerably more sensitive. Final detection was on autoradiography film.

      Read More

      Abcam Scientific Support

      Answered on Oct 03 2003

      Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
      For licensing inquiries, please contact partnerships@abcam.com

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