Overview

  • Product name
    Anti-RBPJK antibody - ChIP Grade
    See all RBPJK primary antibodies
  • Description
    Rabbit polyclonal to RBPJK - ChIP Grade
  • Host species
    Rabbit
  • Tested applications
    Suitable for: IP, ChIP, EMSA, WB, ICC/IFmore details
  • Species reactivity
    Reacts with: Mouse, Human
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 450 to the C-terminus of Human RBPJK.

    Read Abcam's proprietary immunogen policy (Peptides available as ab26893)

  • Positive control
    • This antibody gave a positive signal in Human Testis and Mouse Testis tissue lysates and HeLa whole cell lysate.

Properties

Applications

Our Abpromise guarantee covers the use of ab25949 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
IP Use a concentration of 5 µg/ml.
ChIP Use at an assay dependent concentration. PubMed: 20661424
EMSA Use at an assay dependent concentration. PubMed: 21991380
WB Use a concentration of 1 µg/ml. Detects a band of approximately 56 kDa (predicted molecular weight: 56 kDa).
ICC/IF Use at an assay dependent concentration.

Target

  • Function
    Transcriptional regulator that plays a central role in Notch signaling, a signaling pathway involved in cell-cell communication that regulates a broad spectrum of cell-fate determinations. Acts as a transcriptional repressor when it is not associated with Notch proteins. When associated with some NICD product of Notch proteins (Notch intracellular domain), it acts as a transcriptional activator that activates transcription of Notch target genes. Probably represses or activates transcription via the recruitment of chromatin remodeling complexes containing histone deacetylase or histone acetylase proteins, respectively. Specifically binds to the immunoglobulin kappa-type J segment recombination signal sequence.
  • Sequence similarities
    Belongs to the Su(H) family.
    Contains 1 IPT/TIG domain.
  • Cellular localization
    Nucleus. Cytoplasm. Mainly nuclear, upon interaction with RITA/C12orf52, translocates to the cytoplasm, down-regulating the Notch signaling pathway.
  • Information by UniProt
  • Database links
  • Alternative names
    • AI843960 antibody
    • AOS3 antibody
    • CBF 1 antibody
    • CBF-1 antibody
    • CBF1 antibody
    • csl antibody
    • IGKJRB antibody
    • IGKJRB1 antibody
    • J kappa recombination signal binding protein antibody
    • J kappa-recombination signal-binding protein antibody
    • KBF2 antibody
    • NY REN 30 antigen antibody
    • RBP J antibody
    • RBP J kappa antibody
    • RBP JK antibody
    • RBP-J antibody
    • RBP-J kappa antibody
    • RBP-JK antibody
    • Rbpj antibody
    • RBPJK antibody
    • RBPSUH antibody
    • recombination signal binding protein for immunoglobulin kappa J region antibody
    • Recombining binding protein suppressor of hairless antibody
    • Renal carcinoma antigen NY-REN-30 antibody
    • SUH antibody
    • SUH_HUMAN antibody
    see all

Images

  • All lanes : ab25949 at 1 µg/ml

    Lane 1 : Testis (Human) Tissue Lysate at 20 µg
    Lane 2 : Testis (Mouse) Tissue Lysate at 20 µg
    Lane 3 : HeLa (Human) Whole cell lysate at 10 µg

    Secondary
    All lanes : Anti-Rabbit IgG VHH Single Domain (HRP) at 1/50000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 56 kDa
    Observed band size: 60 kDa
    why is the actual band size different from the predicted?


    Exposure time: 20 minutes


    Abcam recommends using milk as the blocking agent. Abcam welcomes customer feedback and would appreciate any comments regarding this product and the data presented above.

  • Anti-RBPJK antibody - ChIP Grade (ab25949) at 1 µg/ml + Human pancreas tissue lysate - total protein (ab29816) at 20 µg

    Secondary
    Anti-Rabbit IgG VHH Single Domain (HRP) (ab191866) at 1/50000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 56 kDa
    Observed band size: 56 kDa
    Additional bands at: 98 kDa. We are unsure as to the identity of these extra bands.


    Exposure time: 16 minutes


    This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 2% Bovine Serum Albumin before being incubated with ab25949 overnight at 4°C. Antibody binding was detected using an anti-rabbit IgG VHH single domain antibody conjugated to HRP (ab191866), and visualised using ECL development solution ab133406.

  • RBPJK was immunoprecipitated using 0.5mg PANC1 whole cell extract, 5µg of Rabbit polyclonal to RBPJK and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).

    The antibody was incubated under agitation with Protein G beads for 10min, PANC1 whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.

    Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab25949.

    Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).

    Band: 56kDa, non specific bands -50kDa: This extra band is heavy chain contamination; RBPJK

  • All lanes : Anti-RBPJK antibody - ChIP Grade (ab25949) at 1/500 dilution

    Lane 1 : Mouse myoblast C2C12 whole cell lysate at 50 µg
    Lane 2 : Mouse myoblast C2C12 whole cell lysate at 75 µg

    Secondary
    All lanes : HRP-conjugated Goat anti-rabbit at 1/2000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 56 kDa
    Observed band size: 56 kDa


    Exposure time: 1 minute

    See Abreview

References

This product has been referenced in:
  • Szymula A  et al. Epstein-Barr virus nuclear antigen EBNA-LP is essential for transforming naïve B cells, and facilitates recruitment of transcription factors to the viral genome. PLoS Pathog 14:e1006890 (2018). Read more (PubMed: 29462212) »
  • Ting HA  et al. Notch ligand Delta-like 4 induces epigenetic regulation of Treg cell differentiation and function in viral infection. Mucosal Immunol 11:1524-1536 (2018). Read more (PubMed: 30038214) »
See all 15 Publications for this product

Customer reviews and Q&As

1-8 of 8 Abreviews or Q&A

Answer

Thank you for taking the time to confirm this and sending us the qPCR results. The details you have kindly provided will provide us with vital information for our monitoring of product quality. It is regrettable the results have not been successful. When testing our antibodies, our lab uses 5% BSA as a blocking reagent, so I could only recommend switching to this instead of milk for future experiments. For unknown reasons some antibodies give stronger, more specific signals on blots blocked with BSA instead of milk, so doing this may improve the results you are seeing. However, reviewing the details, I am sorry there are no further tips to provide on this occasion to help improve the results in this particular case. I can suggest you have regrettably received a bad vial. I apologise for the inconvenience and 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 details of how you would like to proceed.

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Application
ChIP
Sample
Rat Cell lysate - nuclear (Cardiac fibroblasts)
Negative control
ChIP with rabbit IgG
Specification
Cardiac fibroblasts
Detection step
Real-time PCR
Type
Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
Specification of the cross-linking agent: 1% formaldehyde
Positive control
I didn't use a positive control

Mrs. Tal Golan Lagziel

Verified customer

Submitted Oct 31 2017

Application
Western blot
Loading amount
50 µg
Gel Running Conditions
Reduced Denaturing
Sample
Mouse Cell lysate - whole cell (Mouse myoblast C2C12 cell line)
Specification
Mouse myoblast C2C12 cell line
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Sir or Madam Jkfarmaha

Verified customer

Submitted Oct 29 2013

Answer

I am very pleased to hear you would like to accept our offer and test ab25949 in mouse. Please order the antibody online if you prefer, and have the item deilvered to your Institute. This code will give you 1 free primary antibody before the expiration date. To redeem this offer, please first submit an Abreview for mouse and include this code in the “Additional Comments” section so we know the Abreview is for this promotion. Please remember that submission of the Abreview is sufficient for the discount code to become active.

For more information on how to submit an Abreview, please visit the site: www.abcam.com/Abreviews.

Remember, we publish both positive and negative Abreviews on our datasheets so please submit the results of your tests. The code will be active once the Abreview has been submitted and can be redeemed in one of the following ways:

1) Call to place your order and mention the code to our customer service department;
2) Include the code in your fax order;
3) Place your order on the web and enter the promotional code.

Any feedback that you can provide will be greatly appreciated, whether positive or negative. If you have any further questions, please do not hesitate to contact us. We look forward to receiving your Abreview and wish you luck with your research.

The terms and conditions applicable to this offer can be found here: www.abcam.com/collaborationdiscount.

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Answer

Thank you very much for your interest in ab25949

To our knowledge, ab29549 has not been tested in mouse. Therefore, I can offer a discount off a future purchase if you buy ab25949 now, test it in mouse and submit feedback to us in the form of an Abreview. It doesn’t matter whether the Abreview is positive or negative, we would just really like to receive your feedback. The discount would be to the value of 1 free primary antibody

If you are interested in this offer, please follow these steps:

1. Reply to this e-mail to let me know that you would like to proceed and test ab25949 in mouse. I will then send a discount code. This code must be issued before purchasing ab25949 so please wait for my reply before ordering.

2. Purchase ab25949 either by phone, fax, or online (www.abcam.com).

3. Test it in mouse.

4. Let us know the results, positive or negative, using our Abreview system (this will take about 10 minutes and images are great if you have them!). To find out how to submit an Abreview, please visit: https://www.abcam.com/abreviews.

5. After the review is submitted to us, the discount code becomes active. Simply place your new order by phone, fax, or on the web and mention the discount code. The discount can be redeemed for any primary antibody ordered and the discount code is valid for 4 months after issue.
Please remember that submission of the Abreview is sufficient for the discount code to become active.

We are always pleased to obtain feedback about our products and any information is greatly appreciated! Even if ab25949 turns out to be unsuitable for mouse, you will still receive the discount on your next purchase after your Abreview has been submitted.

Please let me know if you have any questions about this offer and I would be happy to help you further.

The Terms and Conditions of this offer can be found at: www.abcam.com/collaborationdiscount.

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Question
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. As usual if you have any further questions regarding this credit note, please contact the accounts department by email at creditcontrol@abcam.com. The reference number, or credit ID, is xxx. Please refer to this 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.

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Question

Dear Tech Support Team,  Please read the details of customer's complaint. The image is attached.   Kindly advise.    Details  Antibody code: Rb pAB to RBPJK , ab25949 Batch number: GR34586-1 Antibody storage conditions (temperature/reconstitution etc)- -20C, Divided into aliquots. It was never thawed before usage. Description of the problem (high background, wrong band size, more bands, no band etc.)- NO BANDS ! Sample (Species/Cell extract/Nuclear extract/Purified protein/Recombinant protein etc.) Cell Extract from ten different cell lines that express the protein-RBP-Jk.   Sample preparation (Buffer/Protease inhibitors/Heating sample etc.)- RIPA Buffer + Protease inhibitor cocktail  (Sigma) Amount of protein loaded 40 microgram Electrophoresis/Gel conditions (Reducing or Non-reducing gel, % of the gel etc.) SDS-PAGE Transfer and blocking conditions (Buffer/time period, Blocking agent etc.) 100V , 1h Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step) –  According to manufacturer instruction diluted 1:500 and also 1:100 after it didn’t work.  O.N. INCUBATION Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step) - JACKSON Detection method (ECL, ECLPlus etc.) – Commercial ECL kit Positive and negative controls used (please specify)- Different cell lines. Developed the same membrane with different antibodies and it was very successful.   Optimization attempts (problem solving)   How many times have you tried the Western? Twice. Have you run a "No Primary" control? Do you obtain the same results every time? e.g. are the background bands always in the same place?  Yes   Thanks in advance for your reply.  

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Answer

Thank you for taking the time to contact us. I am sorry to hear the customer has had difficulty obtaining satisfactory results from this Anti-RBPJK antibody. I appreciate the time spent in the laboratory and understand the customer's concerns. It is regrettable the results have not been successful. Before deciding how to proceed with this particular case, I would appreciate if you can confirm some details of the procedure that we require for our records: 1) Could you please confirm which cell lines were used (human cell lines)? How did you check that they express RBP-Jk: with RT-PCR? 2) May I ask which blocking reagent was used? 3) Could you please specify the secondary antibody? Can you confirm that it works with other polyclonal antibodies from rabbit? Please be reassured that once we received this information and it confirms that the product is not functioning as cited on the product data sheet, we will be pleased to provide a credit note, free of charge replacement or refund. Thank you for your cooperation. I look forward to hearing from you with the requested details and how the customer would like to proceed.

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: RT°C
Sample
Human Cell (Human umbilical vein endothelial cells)
Specification
Human umbilical vein endothelial cells
Fixative
Paraformaldehyde
Permeabilization
Yes - 1% Triton X-100

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Verified customer

Submitted Apr 14 2010

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