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Anti-Rpt1 抗体 (ab22678)

Submit a review Q&A (2)
Western blot - Anti-Rpt1 antibody (ab22678)

    Key features and details

    • Rabbit polyclonal to Rpt1
    • Suitable for: WB
    • Reacts with: Saccharomyces cerevisiae, Schizosaccharomyces pombe
    • Isotype: IgG

    製品の概要

    • 製品名

      Anti-Rpt1 antibody

    • 製品の詳細

      Rabbit polyclonal to Rpt1

    • 由来種

      Rabbit

    • 特異性

      At optimal dilution there is no cross-reactivity with other members of the YTA family.

    • アプリケーション

      適用あり: WBmore details

    • 種交差性

      交差種: Saccharomyces cerevisiae, Schizosaccharomyces pombe

    • 免疫原

      Recombinant fragment corresponding to Rpt1 aa 1-100.
      Database link: P33299

      Run BLAST with BLAST the sequence with ExPASy Run BLAST with BLAST the sequence with NCBI

    • 特記事項

      Dilute to working strength with phosphate buffered saline pH 7.2-7.4 and 1% normal goat serum (if a goat anti-rabbit IgG linker antibody is to be used).

      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

    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ab22678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
    1/10000 - 1/50000. Detects a band of approximately 52 kDa.
    特記事項
    WB
    1/10000 - 1/50000. Detects a band of approximately 52 kDa.

    ターゲット情報

    • 関連性

      Rpt1 in yeast is one of six ATPases of the 19S regulatory particle of the 26S proteasome involved in the degradation of ubiquitinated substrates; required for optimal CDC20 transcription; interacts with Rpn12p and the E3 ubiquitin-protein ligase Ubr1p. In the yeast S. cerevisiae, 12 different members of a novel gene family of putative ATPases have been identified and characterised. Due to their relationship with TBP1, a human immunodeficiency tat binding protein, they have been designated the YTA family (yeast tat binding analogues). The most outstanding feature of the YTA proteins and their relatives is that they share a region of high similarity which extends to some 300 amino acids in length.

    • 細胞内局在

      Cytoplasmic and Nuclear

    • 参照データベース

      • 別名

        • 26S protease regulatory subunit 7 homolog antibody
        • CIM5 antibody
        • Protein CIM5 antibody
        • TAT-binding homolog 3 antibody
        • YTA3 antibody
        see all

      画像

      • Western blot - Anti-Rpt1 antibody (ab22678)
        Western blot - Anti-Rpt1 antibody (ab22678)
        Anti-Rpt1 antibody (ab22678) at 1/25000 dilution + Purified yeast 26S proteasome preparation

        Developed using the ECL technique.

        Exposure time: 1 minute

      参考文献 (0)

      ab22678 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。

      ab22678 は論文での使用が確認できていません。

      レビューと Q&A

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      レビューを送る 質問を送る

      1-2 of 2 Abreviews or Q&A

      Question

      You report cross-reactivity of the Anti-Rpt1 antibody (ab22678) with
      S.pombe. Have you got any data about this? Did you check cellular
      extracts, or a (partial) proteasome purification as for the image in the
      antibody description (produced using the cerevisiae proteasome)?
      Thanks in advance for your assistance
      and best wishes

      Read More
      Answer

      Please find below some information about the testing conditions:

      Western blot - Single dimension SDS-PAGE of a yeast whole cell extract and purified yeast 26S proteasome followed by Western blotting shows a major band at ˜52kDa. The antibody may commonly be used at dilutions in the range 1:10,000 to 1:50,000. At optimal dilution there is no cross-reactivity with other members of the YTA family.

      Immunoprecipitation - This antibody has not been characterised for use in immunoprecipitation.

      Species reactivity:- This antibody has been demonstrated to react with proteasomes from yeast.

      Reference: Schnall, R., Mannhaupt, G., Stucka, R., Tauer, R., Ehnle, S., Schwarzlose, C., Vetter, I. And Feldmann. Identification of a set of yeast genes coding for a novel family of putative ATPases with high similarity to constituents of the 26S protease complex. Yeast, 10, 1141-1155 (1994).

      Read More

      Question

      Our customer had some problem about these ab22678 antibody. They found the expected bands are faint and there are many vertical straight lines on membrane. We would appreciated if you could help them about the experiment. 1. Order details: a.. Batch number: Lot168293 b.. Abcam order or Purchase order number: ab22678 c.. Antibody storage conditions (temperature/reconstitution etc) 4? 2. Please describe the problem (high background, wrong band size, more bands, no band etc). Expected bands are faint and there are many vertical straight lines on membrane 3. On what material are you testing the antibody in WB? · Species: yeast · Cell extract or Nuclear extract: cell extract protein · Purified protein or Recombinant protein: purified protein (RPT1) 3. The lysate a.. How much protein was loaded: 70-100µg for total extract, 700-1000 ug for IP or GST pulldown a.. What lysis buffer was used: RIPA buffer b.. What protease inhibitors were used: aprotinin, leupeptin, Bestain, Pepstain A, E-64, AEBSF and DTT c.. What loading buffer was used: 6×sample buffer d.. Did you heat the samples: temperature and time: 100?, 5 mins 4. Electrophoresis/Gel conditions/ Transfer conditions a.. Reducing or non reducing gel: reducing gel b.. Gel percentage : 10% SDS-PAGE c.. Transfer conditions: 150 mA, 1 hr (semi-dry) 5. Blocking conditions a.. Buffer: TTBS b.. Blocking agent: milk, BSA, serum, what percentage: 5% nonfat milk+TTBS c.. Incubation time: Overnight d.. Incubation temperature: 4? 6. Primary Antibody a.. Specification (in which species was it raised against): rabbit · At what dilution(s) have you tested this antibody: 1:10000, 1:30000, 1:50000 · What dilution buffer was used: TTBS · Incubation time: 3-4 h · Incubation temperature: RT · What washing steps were done: wash membrane in 10 ml TTBS for 10 min, total 3 times 7. Secondary Antibody a.. Specification (in which species was it raised against)? goat anti rabbit conjugate HRP b.. At what dilution(s) have you tested this antibody: 1:10000 c.. Incubation time 1hr d.. Wash steps: wash membrane in 10 ml TTBS for 10 min, total 3 times e.. Do you know whether the problems you are experiencing come from the secondary? No, because the secondary can work for other primary antibodies 8. Detection method ECl, ECl+, other detection method: ECL 9. Background bands · Have you eliminated the possibility that any background bands could be due to the secondary antibody? (Run a “No primary” control): It is not possible, because the secondary antibody can work for other antibodies without these background · Is the blocking step sufficient? yes · Are your washing steps sufficiently stringent? (Multiple short washes are more effective than fewer longer wash steps) yes · At what size are the bands migrating? Could they be degradation products of your target? · Please provide an image of your blot (as an e-mail attachment, a faxed image is not sufficient) Sample 1 (reprobe anti-GST-Rad23) Sample 2 11. Did you apply positive and negative controls along with the samples? Please specify. When reprobe another antibody, there are no these vertical straight lines 10. Optimization attempts · How many times have you tried the Western? 7-8 times/each student, two students are using this antibody · Do you obtain the same results every time e.g. are background bands always in the same place? yes · What steps have you altered? no

      Read More
      Answer

      Thank you for contacting Abcam regarding ab22678 and for taking the time to provide some details of the customer's Western blot experiments. I am very sorry to hear that he/she is having problem with this particular product. Here I am forwarding my comments/suggestions and hope this will help you customer: - As I understand from your e-mail that customer re-probed the membrane with ant-GST-Rad23. It may well be that the tagging interact with the recognition of the antibody. I can also see on the labelling from the third blot, that the customer carried out IP (Flag-IP) but this antibody has only been tested for Western blot application but not for immunoprecipitation (IP). - Multiple bands could be due to degradation of the proteins so it is very important to make sure that the freshly prepared lysis buffer is used for sample preparation. - Block the membrane for shorter time for 1 hr instead of overnight at 4oC. Too long blocking can also result in non-specific signals and fainted bands. I would like to emphasize that since this antibody was purchased in September 2006 - almost 5 months - I regret to inform you that the Abcam’s guarantee (free of charge replacement or credit note) does not apply for this purchase since we have no control how the antibody was stored. I am very sorry but I can’t be any of further assistance.

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