Anti-Apc11 antibody (ab44708)

Submit a review Q&A (4)References (1)

Overview

  • Product name
    Anti-Apc11 antibody
    See all Apc11 primary antibodies
  • Description
    Rabbit polyclonal to Apc11
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WBmore details
  • Species reactivity
    Reacts with: Mouse, Human
  • Immunogen

    Full length protein corresponding to Human Apc11.

  • Positive control
    • HeLa, NIH-3T3 cells

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage buffer
    pH: 7.40
    Preservative: 0.1% Sodium azide
    Constituent: 0.01% PBS
  • Concentration information loading...
  • Purity
    Protein G purified
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab44708 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
WB 1/400. Predicted molecular weight: 10 kDa.

Target

  • Function
    Component of the anaphase promoting complex/cyclosome (APC/C), a cell cycle-regulated E3 ubiquitin ligase that controls progression through mitosis and the G1 phase of the cell cycle. The APC/C complex acts by mediating ubiquitination and subsequent degradation of target proteins: it mainly mediates the formation of 'Lys-11'-linked polyubiquitin chains and, to a lower extent, the formation of 'Lys-48'- and 'Lys-63'-linked polyubiquitin chains. May recruit the E2 ubiquitin-conjugating enzymes to the complex.
  • Tissue specificity
    Expressed at high levels in skeletal muscle and heart; in moderate levels in brain, kidney, and liver; and at low levels in colon, thymus, spleen, small intestine, placenta, lung and peripheral blood leukocyte.
  • Pathway
    Protein modification; protein ubiquitination.
  • Sequence similarities
    Belongs to the RING-box family.
    Contains 1 RING-type zinc finger.
  • Domain
    The RING-type zinc finger domain coordinates an additional third zinc ion.
  • Post-translational
    modifications
    Auto-ubiquitinated.
  • Cellular localization
    Cytoplasm. Nucleus.
  • Information by UniProt
  • Database links
    • Alternative names
      • ANAPC 11 antibody
      • ANAPC11 antibody
      • Anaphase promoting complex subunit 11 (yeast APC11 homolog) antibody
      • Anaphase promoting complex subunit 11 antibody
      • Anaphase promoting complex subunit 11 homolog (yeast) antibody
      • Anaphase promoting complex subunit 11 homolog antibody
      • Anaphase-promoting complex subunit 11 antibody
      • Apc 11 antibody
      • Apc 11p antibody
      • APC11 anaphase promoting complex subunit 11 homolog (yeast) antibody
      • APC11 anaphase promoting complex subunit 11 homolog antibody
      • APC11 antibody
      • APC11_HUMAN antibody
      • Apc11p antibody
      • Cyclosome subunit 11 antibody
      • Hepatocellular carcinoma associated RING finger protein antibody
      • Hepatocellular carcinoma-associated RING finger protein antibody
      • HSPC 214 antibody
      • HSPC214 antibody
      • MGC882 antibody
      • Yeast APC 11 homolog antibody
      • Yeast APC11 homolog antibody
      see all

    References

    This product has been referenced in:
    See 1 Publication for this product

    Publishing research using ab44708? Please let us know so that we can cite the reference in this datasheet.

    Customer reviews and Q&As

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    1-4 of 4 Abreviews or Q&A

    Question

    Dear Keith, Thanks for your reply. I really don¹t think its a problem with the secondary. I have used this secondary for 20 years now and it works really well. I have used it with many other antibodies in all of the buffers that I detailed in my enquiry and it doesn¹t give me this problem. Also, since I have no bands at all in marvel (this is what I use for almost every antibody I use apart from a few phospho-antibodies) I don¹t think that washing for longer will help (again, these are conditions that I use for all my antibodies, although I appreciate that some antibodies may require alternative conditions). I could wash for longer when I use BSA but it is possible to see through the background and there are no bands apparent there either. I will you send images of the blots later today but my "specific issue" with this antibody is that it really doesn¹t appear to work. Do you have any data to show that it does, particularly for detection of endogenous protein? Thanks for your time, Lindsey Lindsey A Allan PhD. Division of Cancer Research Medical Research Institute University of Dundee Level 5 Ninewells Hospital & Medical School Ninewells Drive Dundee DD1 9SY Scotland, UK Tel: +44(0)1382 496659 e-mail: l.a.allan@dundee.ac.uk On 10/01/2012 22:44, "technical@abcam.com" wrote: > > > Dear Dr Allan, > >We have an answer to your inquiry: >Thank you for contacting us. > >I am sorry to hear you are experiencing difficulties with one of our >products. We take product complaints very seriously, and investigate >every product that we feel may not be performing correctly. > >Have you tried tittering the secondary antibody for use with this assay? >Your experience with no band or black blots may be a result of your >secondary antibody. Often 'black blot' issues are result of secondary >antibody reacting with the different blocking solutions. I would >recommend more and longer washes with TBST as well. > >Would you be able to send me images of your blots? Seeing these may help >to solve your specific issues. > >I look forward to receiving your reply. Please let me know if you have >any questions. >Help us improve our service. Rate your experience with us today. >https://www.abcam.com/index.html?pageConfig=technicalSurvey&intCCEID=341412 >9 > >Your original inquiry to Abcam: >------------------------------------------- > > LOT NUMBER GR42267-1 ORDER NUMBER BRI12016 DESCRIPTION OF THE PROBLEM No >signal or completely black depending upon buffer used (see primary >antibody section) SAMPLE Human cell lysate from U2OS cells and DLD-1 >cells. PRIMARY ANTIBODY Primary antibody diluted 1/400 in 5% marvel, 5% >BSA or Reliablot all in TBS-0.2% Tween. Incubated 4C overnight Washes: >TBS-0.2% Tween, 1x 10min, 2x 5min, RT I have also tried combining a BSA >block with a marvel primary. In all cases, if i use marvel to block the >blot is blank below 30K where there are some faint bands. If i use BSA at >either stage or Reliablot the blot is totally black. DETECTION METHOD ECL >made in our lab. This is the only ECL we use and it is sensitive enough >to be used for the many phospho-antibodies we use. It is used with X-ray >film. POSITIVE AND NEGATIVE CONTROLS USED I have no specific >positive/negative controls. These cell lines have functional APC-C so >they must express APC11. I have tried to knock APC 11 down using siRNA >(which was why i wanted the antibody in the first place, to confirm the >degree of knockdown). I am using a Dharmacon Smartpool plus for this and >i have never had one of these not work. However, since this antibody is >not picking up any bands in the control cells there is no way to know if >the knockdown is working! ANTIBODY STORAGE CONDITIONS +4C SAMPLE >PREPARATION Cell lysate prepared in RIPA buffer containing protease >inhibitor cocktail. 4x SDS-loading buffer containing 2-ME (final conc = >5%) added prior to heating to 100C, 4min. AMOUNT OF PROTEIN LOADED Up to >20ug on 15-well, 1mm SDS-PAGE ELECTROPHORESIS/GEL CONDITIONS 15% reducing >SDS-PAGE, run until 10K marker roughly in middle of resolving gel. >TRANSFER AND BLOCKING CONDITIONS Transfer: semi-dry transfer, 12V, 30min >onto 0.45uM Hybond ECL nitrocellulose. Then tried small pore 0.2uM Hybond >ECL nitrocellulose. Transfer buffer: 20mM Tris150mM Glycine0.1% SDS >Blocking agent: 5% marvel, 5% BSA or Reliablot all in TBS-0.2% Tween, all >for 60min, RT SECONDARY ANTIBODY Secondary antibody: Biorad >anti-rabbit-HRP diluted 1/5000 in 5% marvel in TBS-0.2% Tween, 60min, RT >Washes:TBS-0.2% Tween, 1x 10min, 2x 5min, RT HOW MANY TIMES HAVE YOU >TRIED THE APPLICATION? 4 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU >OBTAIN THE SAME RESULTS EVERY TIME? No WHAT STEPS HAVE YOU ALTERED? As >detailed above: 1. pore size of membrane 2. blocking buffer 3. primary >antibody dilution buffer ADDITIONAL NOTES Can you let me know if you have >any specific protocols for using this antibody for western blotting >please. If not, would you consider sending an alternative antibody for me >to test please? None of the APC11 antibodies seem to have any AbReview >and there is only one with a WB photo but that is of recombinant protein. >Thank you, Lindsey >------------------------------------------- > > >Best regards, >Keith > >Keith Beadle >Scientific Support Specialist >Abcam Inc. >www.abcam.com > > >Abcam Customer Services and Technical Support Team > >www.abcam.com/technical >[CCE3414129] > > The University of Dundee is a registered Scottish Charity, No: SC015096

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    Answer

    Thank you for contacting us. I have spoken with the lab about this issue.They have provided me with the following information. The conditions used in-house are essentially the same as those in your protocol, however, we are unable to locate anything specifically for it working in endogenous proteins. I would like to offer a free of charge replacement antibody to you. We may send another vial of ab44708 or another antibody to the target. Please let me know which you prefer, I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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    Question

    Hello, Would you provide me with an update on when I can expect this antibody please? The last estimate I had was that it would be shipped around 25th Jan but that I would receive an e-mail when it was shipped. I have not had an e-mail. It would be really helpful to have it ASAP - I have been waiting for a successful APC11 blot since November when I originally ordered the one that subsequently failed to work. Its far from ideal in research when things take so long.

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    Answer

    Thank you for contacting us.

    It appears that this product is overdue and is not expected to ship for 1 -2 weeks. I unable to find any further information as to the cause of the delay. I am very sorry for the delay. Please let me know if there is anything I may do for you.

    I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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    Question

    Hello, Thanks for your reply. I am pleased that you will provide a different antibody FOC. From your website, there don’t appear to be any antibodies that are known to work on endogenous protein. However, one of them does show a western with recombinant protein. Unless you can advise otherwise, this may be the best one to try next: Anti-APC11 antibody, ab57158. Of course I will let you know how I get on with it. Many thanks for your help,

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    Answer

    Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products. I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement with the order number XXXXX. This product is currently not in stock but is scheduled for delivery around 25 Jan 2012. A notification will be send when it ships. To check the status of the order please contact our Customer Service team and reference this number. Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know. I would like to know how this replacement performs for you and I wish you the best of luck with your research.

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    Question

    DESCRIPTION OF THE PROBLEM No signal or completely black depending upon buffer used (see primary antibody section) SAMPLE Human cell lysate from U2OS cells and DLD-1 cells. PRIMARY ANTIBODY Primary antibody diluted 1/400 in 5% marvel, 5% BSA or Reliablot all in TBS-0.2% Tween. Incubated 4C overnight Washes: TBS-0.2% Tween, 1x 10min, 2x 5min, RT I have also tried combining a BSA block with a marvel primary. In all cases, if i use marvel to block the blot is blank below 30K where there are some faint bands. If i use BSA at either stage or Reliablot the blot is totally black. DETECTION METHOD ECL made in our lab. This is the only ECL we use and it is sensitive enough to be used for the many phospho-antibodies we use. It is used with X-ray film. POSITIVE AND NEGATIVE CONTROLS USED I have no specific positive/negative controls. These cell lines have functional APC-C so they must express APC11. I have tried to knock APC 11 down using siRNA (which was why i wanted the antibody in the first place, to confirm the degree of knockdown). I am using a Dharmacon Smartpool plus for this and i have never had one of these not work. However, since this antibody is not picking up any bands in the control cells there is no way to know if the knockdown is working! ANTIBODY STORAGE CONDITIONS +4C SAMPLE PREPARATION Cell lysate prepared in RIPA buffer containing protease inhibitor cocktail. 4x SDS-loading buffer containing 2-ME (final conc = 5%) added prior to heating to 100C, 4min. AMOUNT OF PROTEIN LOADED Up to 20ug on 15-well, 1mm SDS-PAGE ELECTROPHORESIS/GEL CONDITIONS 15% reducing SDS-PAGE, run until 10K marker roughly in middle of resolving gel. TRANSFER AND BLOCKING CONDITIONS Transfer: semi-dry transfer, 12V, 30min onto 0.45uM Hybond ECL nitrocellulose. Then tried small pore 0.2uM Hybond ECL nitrocellulose. Transfer buffer: 20mM Tris150mM Glycine0.1% SDS Blocking agent: 5% marvel, 5% BSA or Reliablot all in TBS-0.2% Tween, all for 60min, RT SECONDARY ANTIBODY Secondary antibody: Biorad anti-rabbit-HRP diluted 1/5000 in 5% marvel in TBS-0.2% Tween, 60min, RT Washes:TBS-0.2% Tween, 1x 10min, 2x 5min, RT HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 4 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? No WHAT STEPS HAVE YOU ALTERED? As detailed above: 1. pore size of membrane 2. blocking buffer 3. primary antibody dilution buffer ADDITIONAL NOTES Can you let me know if you have any specific protocols for using this antibody for western blotting please. If not, would you consider sending an alternative antibody for me to test please? None of the APC11 antibodies seem to have any AbReview and there is only one with a WB photo but that is of recombinant protein. Thank you,

    Read More
    Answer

    Thank you for contacting us. I am sorry to hear you are experiencing difficulties with one of our products. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly. Have you tried tittering the secondary antibody for use with this assay? Your experience with no band or black blots may be a result of your secondary antibody. Often 'black blot' issues are result of secondary antibody reacting with the different blocking solutions. I would recommend more and longer washes with TBST as well. Would you be able to send me images of your blots? Seeing these may help to solve your specific issues. I look forward to receiving your reply. Please let me know if you have any questions.

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