Overview

  • Product name
    Anti-CREB antibody [E306]
    See all CREB primary antibodies
  • Description
    Rabbit monoclonal [E306] to CREB
  • Host species
    Rabbit
  • Specificity
    This antibody recognizes CREB, and can also recognize the splice isoform of CREB.
  • Tested applications
    Suitable for: WB, ICC/IF, IHC-P, IPmore details
    Unsuitable for: Flow Cyt
  • Species reactivity
    Reacts with: Mouse, Rat, Human
    Predicted to work with: Cow
  • Immunogen

    Synthetic peptide within Human CREB aa 250-350. The exact sequence is proprietary.

  • Positive control
    • ICC/IF: PC 12 cells. WB: A431 cell lysate. IHC-P: Prostatic carcinoma tissue.
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab32515 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 1/500 - 1/1000. Detects a band of approximately 40 kDa (predicted molecular weight: 37 kDa).
ICC/IF 1/250 - 1/500.
IHC-P Use at an assay dependent concentration.
IP 1/80.
  • Application notes
    Is unsuitable for Flow Cyt.
  • Target

    • Function
      This protein binds the cAMP response element (CRE), a sequence present in many viral and cellular promoters. CREB stimulates transcription on binding to the CRE. Transcription activation is enhanced by the TORC coactivators which act independently of Ser-133 phosphorylation. Implicated in synchronization of circadian rhythmicity.
    • Involvement in disease
      Defects in CREB1 may be a cause of angiomatoid fibrous histiocytoma (AFH) [MIM:612160]. A distinct variant of malignant fibrous histiocytoma that typically occurs in children and adolescents and is manifest by nodular subcutaneous growth. Characteristic microscopic features include lobulated sheets of histiocyte-like cells intimately associated with areas of hemorrhage and cystic pseudovascular spaces, as well as a striking cuffing of inflammatory cells, mimicking a lymph node metastasis. Note=A chromosomal aberration involving CREB1 is found in a patient with angiomatoid fibrous histiocytoma. Translocation t(2;22)(q33;q12) with CREB1 generates a EWSR1/CREB1 fusion gene that is most common genetic abnormality in this tumor type.
    • Sequence similarities
      Belongs to the bZIP family.
      Contains 1 bZIP domain.
      Contains 1 KID (kinase-inducible) domain.
    • Post-translational
      modifications
      Stimulated by phosphorylation. Phosphorylation of both Ser-133 and Ser-142 in the SCN regulates the activity of CREB and participates in circadian rhythm generation. Phosphorylation of Ser-133 allows CREBBP binding (By similarity). Phosphorylated upon DNA damage, probably by ATM or ATR.
      Sumoylated by SUMO1. Sumoylation on Lys-304, but not on Lys-285, is required for nuclear localization of this protein. Sumoylation is enhanced under hypoxia, promoting nuclear localization and stabilization.
    • Cellular localization
      Nucleus.
    • Information by UniProt
    • Database links
    • Alternative names
      • Active transcription factor CREB antibody
      • cAMP response element binding protein 1 antibody
      • cAMP response element binding protein antibody
      • cAMP responsive element binding protein 1 antibody
      • cAMP-responsive element-binding protein 1 antibody
      • CREB antibody
      • CREB-1 antibody
      • CREB1 antibody
      • CREB1_HUMAN antibody
      • Cyclic AMP-responsive element-binding protein 1 antibody
      • MGC9284 antibody
      • OTTHUMP00000163864 antibody
      • OTTHUMP00000163865 antibody
      • OTTHUMP00000206660 antibody
      • OTTHUMP00000206662 antibody
      • OTTHUMP00000206667 antibody
      • Transactivator protein antibody
      see all

    Images

    • All lanes : Anti-CREB antibody [E306] (ab32515) at 0.03 µg/ml

      Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg
      Lane 2 : A431 (Human epidermoid carcinoma epithelial cell) whole cell lysate at 20 µg
      Lane 3 : PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate at 20 µg
      Lane 4 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

      Predicted band size: 37 kDa
      Observed band size: 40 kDa
      why is the actual band size different from the predicted?


      Exposure time: 15 seconds


      Blocking and diluting buffer: 5% NFDM/TBST.

    • Lane 1: Wild-type HAP1 cell lysate (20 µg)
      Lane 2: CREB knockout HAP1 cell lysate (20 µg)
      Lane 3: HeLa cell lysate (20 µg)
      Lane 4: A431 cell lysate(20 µg)

      Lanes 1 - 4: Merged signal (red and green). Green - ab32515 observed at 44 kDa. Red - loading control, ab18058, observed at 214 kDa.

      ab32515 was shown to recognize CREB in wild-type HAP1 cells along with additional cross-reactive bands. No band was observed when CREB knockout samples examined. Wild-type and knockout samples were subjected to SDS-PAGE. ab32515 and ab18058 (loading control to Vinculin) diluted to 1/500 and 1/10000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

    • ab32515 staining CREB in HeLa cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde and permeabilized with 0.5% Triton X-100 in PBS. Samples were incubated with primary antibody (1/500 in PBS) for 1 hour at 22°C. ab150081, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG polyclonal (1/200) was used as the secondary antibody. Counterstained with DAPI.

      See Abreview

    • Ab32515, at a 1/500 dilution, staining CREB in paraffin embedded prostatic carcinoma tissue sections by Immunohistochemistry.

    References

    This product has been referenced in:
    • Wu XP  et al. MicroRNA-365 alleviates morphine analgesic tolerance via the inactivation of the ERK/CREB signaling pathway by negatively targeting ß-arrestin2. J Biomed Sci 25:10 (2018). WB . Read more (PubMed: 29415719) »
    • Li X  et al. Cisplatin Enhances Hepatitis B Virus Replication and PGC-1a Expression through Endoplasmic Reticulum Stress. Sci Rep 8:3496 (2018). Read more (PubMed: 29472690) »
    See all 29 Publications for this product

    Customer reviews and Q&As

    1-8 of 8 Abreviews or Q&A

    Application
    Western blot
    Sample
    Zebrafish Tissue lysate - whole (Testicle)
    Gel Running Conditions
    Reduced Denaturing (12)
    Loading amount
    5 µg
    Specification
    Testicle
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

    Abcam user community

    Verified customer

    Submitted Apr 17 2018

    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (Hela)
    Gel Running Conditions
    Reduced Denaturing (12%)
    Loading amount
    10 µg
    Specification
    Hela
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C

    Dr. Lin Zhu

    Verified customer

    Submitted Sep 04 2017

    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (HeLa)
    Specification
    HeLa
    Permeabilization
    Yes - 0.5% Triton-X100 in PBS
    Fixative
    Paraformaldehyde

    Dr. Kirk Mcmanus

    Verified customer

    Submitted Nov 05 2014

    Answer

    Thank you for this additional information. I would recommend testing under reducing and denaturing conditions rather than a native gel, as this is how we have tested this product. Otherwise your protocol seems good. Please let me know if you are still having difficulty with this product. Normally our guarantee for 6 months from purchase, but if it was stored properly at -20C in aliquots then we can make an exception as a one time courtesy.

    Read More

    Answer

    Thank you for confirming this information and for your help and cooperation with this case.

    As requested, I have asked our accounting department to issue you with a credit note. This can then be redeemed against the invoice of a future order.

    Credit ID: 20123

    As usual if you have any further questions regarding this credit note, please contact the accounts department by email at creditcontrol@abcam.com. Please refer to the credit ID number in any correspondence with the accounting department.

    I would like to wish the customer good luck with their research. The technical team is always at your service, should you require further expert advice. Have a good day!

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    Question
    Answer

    Thank you for taking the time to complete our questionnaire and contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody.

    I appreciate the time you have spent in the laboratory and understand your concerns. It is regrettable the results have not been successful. Reviewing the details, I am sorry there are only a few tips to provide on this occasion to help improve the results. However, I can recommend you may like to consider trying the following options:

    General: 1.

    We would always recommend to optimize conditions first on less precious samples.

    ICC:

    1. We recommend not to mix blocking agents in one experiment as this can affect the results. Also, we would recommend to use 2%, 5% or 10% serum. 30% is quite high. Or you can use 2% or 5% BSA.

    2. Could you confirm how long was the fixation in PFA? I suggest to try 10 minutes only if this has not already been tried.

    3. I can recommend to fix and then permeabilize. Don’t do both at once.

    WB:

    1. I can recommend to check the amount of protein loaded for the pre embryo samples.

    2. It is unusual that the T47D Cell sample is giving a band at the correct size, but the pre embryo sample is not. I can suggest to review the preparation procedure. Also, I have tried a literature to ensure there is nothing different that would be expected from these sample. I have been unable to find anything, but can suggest you may like to try a more extensive search as this may provide an explanation.

    3. I can recommend to try heating to 70oC only when reducing and denaturing as this will help to prevent formation of protein aggregates which could affect the results.

    Alternatively, or if these tips do not work, I am pleased to offer you a free of charge replacement or credit note in compensation.

    Thank you for your cooperation. I look forward to hearing from you with results from the next experiment and/or details of how you would like to proceed.

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    Question
    Answer

    Thank you for contacting Abcam.



    The concentration for ab32515, lot #GR50483-2, is**********.

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

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    Answer

    Codes offre de test :
    ab32515 : *****
    ab32096 : *****
    ab7217 : *****
    ab12135 : *****
    ab1791 : *****

    Date d’expiration : JJ MM AAAA

    Je suis ravi que vous souhaitiez participer ànotre offre et tester les anticorps listés ci-dessusavec le robot Simon de Protein Simple.Ces codes de réductionsont utilisables sous 120 jours et pour l’achat d’autres anticorps primaires.

    Vous devez dans un premier temps soumettre une Abreview avec vos résultats enspécifiant le code promotionnel correspondant à l'anticorps testé. Ce code sera ainsi activé et vous pourrez par la suite contacter notre service clientèle par téléphone pour passer votre prochaine commande, en vous munissant des informations suivantes : code de réduction, numéro de commande de l'anticorps testé.

    Concernantles produits pour lesquels vousavez déjà obtenu des résultats, merci dem'envoyer vos données, en échange nous vous attribueronsdes"Abpoints" utilisables pour des remisessur vos commandes ouen bons d'achatAmazon.

    Je reste à votre disposition pour plus d’informations concernant cette offre promotionnelle. Dans l’attente de recevoir votre Abreview je vous souhaite unebonne continuation dans vos recherches.

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