Overview

  • Product name
    Anti-CREB (phospho S129 + S133) antibody
    See all CREB primary antibodies
  • Description
    Rabbit polyclonal to CREB (phospho S129 + S133)
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WBmore details
  • Species reactivity
    Reacts with: Mouse, Human
    Predicted to work with: Rat
  • Immunogen

    Synthetic peptide (Human).

  • Positive control
    • NIH3T3 cells with and without PDGF stimulation; Y-1 cells.

Properties

Applications

Our Abpromise guarantee covers the use of ab10564 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 1/1000. Detects a band of approximately 43 kDa (predicted molecular weight: 37 kDa).

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

  • TPeptide Competition and Phosphatase Treatment: Lysates prepared from NIH3T3 cells stimulated with PDGF were resolved by SDS-PAGE on a 10% polyacrylamide gel and transferred to PVDF. Membranes were either left untreated (1-5) or treated with lambda phosphatase (6), blocked with a 5% BSA-TBST buffer for one hour at room temperature, and incubated with ab10564 antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1, 2, 6), the non-phosphopeptide corresponding to the immunogen (3), a generic phosphoserine-containing peptide (4), or, the phosphopeptide immunogen (5). After washing, membranes were incubated with goat F(ab’)2 anti-rabbit IgG HRP conjugate and bands were detected using the chemilumiescence. The data show that the peptide corresponding to ab10564 blocks the antibody signal. The data also show that phosphatase stripping eliminates the signal, verifying that the antibody is phospho-specific. TPeptide Competition and
  • Immunohistochemistry (PFA perfusion fixed frozen section) staining of CREB (phospho S129 + S133) in the mouse cortex. Samples were fixed with paraformaldehyde and ab10564 was used at 1/1000 for 18 hours at 25ºC. An Alexa Fluor®568 conjugated donkey anti-Rabbit secondary was used at 1/1000.

    See Abreview

References

This product has been referenced in:
  • Peres RS  et al. TGF-ß signalling defect is linked to low CD39 expression on regulatory T cells and methotrexate resistance in rheumatoid arthritis. J Autoimmun 90:49-58 (2018). WB . Read more (PubMed: 29426578) »
  • Su Y  et al. Receptor desensitization and blockade of the suppressive effects of prostaglandin E(2) and adenosine on the cytotoxic activity of human melanoma-infiltrating T lymphocytes. Cancer Immunol Immunother 60:111-22 (2011). WB ; Human . Read more (PubMed: 20960188) »
See all 3 Publications for this product

Customer reviews and Q&As

1-8 of 8 Abreviews or Q&A

Application
Western blot
Sample
Rat Tissue lysate - whole (Hippocampus)
Gel Running Conditions
Non-reduced Denaturing (15% SDS-PAGE)
Loading amount
25 µg
Treatment
RIPA & 1% Protease inhibitor for 15min on ice
Specification
Hippocampus
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Oct 10 2018

Application
Western blot
Loading amount
150 µg
Gel Running Conditions
Reduced Denaturing (gel 10% SDS-PAGE)
Sample
Mouse Tissue lysate - whole (Brain)
Specification
Brain
Treatment
Forskoline 1mg/kg i.p.
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Dr. Karine Thibault

Verified customer

Submitted Nov 04 2014

Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Antigen retrieval step
None
Sample
Mouse Tissue sections (Brain)
Specification
Brain
Permeabilization
No
Fixative
Paraformaldehyde

Dr. Karine Thibault

Verified customer

Submitted Aug 19 2014

Answer

Thank you for contacting us. Because we carry over 70,000 products, it isn't feasible for us to keep small sample sizes of our products.

We are happy to reassure our customers that all of our products are covered by our Abpromise, which guarantees that the product will work in the applications and species specified on the datasheet, or we will offer a replacement, credit, or refund within 6 months of purchase.

If the product is to be used in an untested species or application, you may be eligible for our testing discount program if the antibody has not yet been purchased. Please contact our Scientific Support team by replying to this email prior to purchase for more information.

Otherwise, we like to encourage all of our customers to submit an Abreview via the online product datasheet. We always appreciate customer feedback, whether positive or negative, and we make all product information available to researchers. Plus, each Abreview earns Abpoints that can be used for discounts on future purchases or rewards such as Amazon.com gift certificates.

To find out more about our Abreview system, please see the following link:

https://www.abcam.com/abreviews

I hope this information is helpful. Please do not hesitate to contact us again with any other questions.

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Answer

Thank you for getting back to me. Yes, it seems that this product might be binding specifically. In which case, I am more than happy to offer you a replacement vial of your choice, ab47373. Please confirm your shipping address/purchasing agent so that I can immediately send you the replacement vial. I hope to receive your confirmation soon. Have a nice day.

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Question

BATCH NUMBER 212697 ORDER NUMBER 227372 DESCRIPTION OF THE PROBLEM It appears to be detecting total CREB in western blots. We are studying CREB phosphorylation on serine133 and in unstimulated cells there is no background with serine133 phospho-CREB Ab. We can see very good induction of serine133 phosphorylation in stimulated cells. When we use the serine 129/133 diphospho-Ab we see a band in all lanes that looks like the signal we get with the total CREB Ab. SAMPLE We are using IEC-18 cells which are epithelial cells derived from the rat ileum. PRIMARY ANTIBODY We incubate overnight at 4 C primary Ab in TBST with azide followed by 3 TBST washes. DETECTION METHOD ECL, GE HealthCare POSITIVE AND NEGATIVE CONTROLS USED We don't have any positive controls for the dual phosphoCREB. ANTIBODY STORAGE CONDITIONS -20 C, aliquoted SAMPLE PREPARATION We treat the cells and lyse them on the dish with SDS-lysis buffer. We heat the samples 95 C for 10 min. ELECTROPHORESIS/GEL CONDITIONS We are using either Pierce or Invitrogen gradient PAGE 4-20%. TRANSFER AND BLOCKING CONDITIONS We electroblot onto PVDF paper using the invitrogen system. We block for 1 hr with either BSA or milk 5% in TBST. SECONDARY ANTIBODY We use either GE Healthcare or Santa Cruz secondary HRP linked Abs, 1:2500-1:6000 for 1 hr in TBST. 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? Yes ADDITIONAL NOTES I would like to try the Ab to phospho-CREB serine129 #AB47373.

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Answer

Thank you for your patience. I am sorry to hear that you are experiencing difficulties with this product ab10564 in western blot. Often it is possible to make suggestions that help resolve problems. We will happily offer technical support and in the event that a product is not functioning in the applications cited on the product data sheet (and the problem has been reported within 120 days of purchase), and if it appears that the antibody is at fault, a replacement/credit note/refund will be offered. I have looked through the protocols you used and have a few questions and suggestions that might help you resolve the problem. If the suggestions do not prove to be helpful, would you please be so kind to confirm the following items in order to help me better understand the cause of the problem. - How much protein did you used? - What was the dilution for the primary antibody? - Can you please confirm that you were able to detect the bands at the specific molecular weight (43kDa)? - When you said you detected bands in all lanes, were there any bands in negative controls as well? If you can also provide an image that would certainly help a lot. - One possible reason why you are observing bands in all the lanes is maybe because the secondary antibody is binding non-specifically. Please try running a no-primary antibody experiment to eliminate this possibility. After doing this, if you still get bands, then it means that your secondary is the problem. However, if you do not get any bands, then the reason is clear that our primary antibody is at fault. Please get back to me once you have confirmed the above matters. If this product is indeed not performing as stated on the datasheet, I am more than happy to offer you a free replacement. I look forward to hearing from you again in order to resolve the matter. Should you require anything further, please do not hesitate to contact me.

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Answer

Thank you for your enquiry. I am sorry that my response has taken so long an answer has not been forthcoming form the lab. The only antibody that I have details of where it has been tested for reactivity against CREM and ATF is ab30651. This antibody has been shown to react with CREB phospho-Ser133 and related protein ATF-1 when phosphorylated on S63. It does not react with any non-phospho-forms of CREB and ATF-1. Furthermore it does not react with other types of CREB and/or ATF related proteins. I hope this information helps, please do not hesitate to contact us if you need any more advice or information.

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Answer

Thank you for your enquiry. To our knowledge this antibody has yet to be tested for cross reactivity with pS133-CREB. At the moment, all the information we have available is that which is listed on the datasheet. If you choose to test this antibody for cross reactivity with pS133-CREB, please let us know how you get on. In return, we will reward you with Abcam loyalty points.

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