Recombinant
RabMAb

Recombinant Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)

Reviews (2)Q&A (1)References (50)

Overview

  • Product name

    Anti-c-Jun antibody [E254] - ChIP Grade
    See all c-Jun primary antibodies

  • Description

    Rabbit monoclonal [E254] to c-Jun - ChIP Grade

  • Host species

    Rabbit

  • Specificity

    The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

  • Tested applications

    Suitable for: WB, IHC-P, IP, ICC/IF, ChIPmore details
    Unsuitable for: Flow Cyt

  • Species reactivity

    Reacts with: Mouse, Rat, Human
    Predicted to work with: Pig

  • Immunogen

    Synthetic peptide within Human c-Jun aa 1-100 (N terminal). The exact sequence is proprietary.

  • Epitope

    ab32137 reacts with an epitope located in the N terminal region of c-Jun.

  • Positive control

    • WB: NIH 3T3 and HeLa cell lysate. ICC/IF HeLa and NIH/3T3 cells. IHC-P: Human lung carcinoma tissue IP: HEK-293 and NIH3T3 cells.

  • General notes

    A trial size is available to purchase for this antibody.

     

     

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

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

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

Properties

Applications

Our Abpromise guarantee covers the use of ab32137 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 - 1/10000. Predicted molecular weight: 36 kDa.
IHC-P 1/100 - 1/250. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.
IP 1/60.
ICC/IF 1/50 - 1/250.
ChIP Use at an assay dependent concentration.
  • Application notes
    Is unsuitable for Flow Cyt.

    • Target

    • Function

      Transcription factor that recognizes and binds to the enhancer heptamer motif 5'-TGA[CG]TCA-3'. Promotes activity of NR5A1 when phosphorylated by HIPK3 leading to increased steroidogenic gene expression upon cAMP signaling pathway stimulation. Involved in activated KRAS-mediated transcriptional activation of USP28 in colorectal cancer (CRC) cells (PubMed:24623306). Binds to the USP28 promoter in colorectal cancer (CRC) cells (PubMed:24623306).

    • Sequence similarities

      Belongs to the bZIP family. Jun subfamily.
      Contains 1 bZIP (basic-leucine zipper) domain.

    • Post-translational
      modifications

      Ubiquitinated by the SCF(FBXW7), leading to its degradation. Ubiquitination takes place following phosphorylation, that promotes interaction with FBXW7.
      Phosphorylated by CaMK4 and PRKDC; phosphorylation enhances the transcriptional activity. Phosphorylated by HIPK3. Phosphorylated by DYRK2 at Ser-243; this primes the protein for subsequent phosphorylation by GSK3B at Thr-239. Phosphorylated at Thr-239, Ser-243 and Ser-249 by GSK3B; phosphorylation reduces its ability to bind DNA. Phosphorylated by PAK2 at Thr-2, Thr-8, Thr-89, Thr-93 and Thr-286 thereby promoting JUN-mediated cell proliferation and transformation. Phosphorylated by PLK3 following hypoxia or UV irradiation, leading to increase DNA-binding activity.
      Acetylated at Lys-271 by EP300.

    • Cellular localization

      Nucleus.
    • Information by UniProt

    • Database links

      see all

    • Alternative names

      • Activator protein 1 antibody
      • AP 1 antibody
      • AP-1 antibody
      • AP1 antibody
      • cJun antibody
      • Enhancer Binding Protein AP1 antibody
      • Jun Activation Domain Binding Protein antibody
      • JUN antibody
      • Jun oncogene antibody
      • JUN protein antibody
      • Jun proto oncogene antibody
      • JUN_HUMAN antibody
      • JUNC antibody
      • Oncogene JUN antibody
      • p39 antibody
      • Proto oncogene c jun antibody
      • Proto oncogene cJun antibody
      • Proto-oncogene c-jun antibody
      • Transcription Factor AP 1 antibody
      • Transcription factor AP-1 antibody
      • Transcription Factor AP1 antibody
      • V jun avian sarcoma virus 17 oncogene homolog antibody
      • V jun sarcoma virus 17 oncogene homolog (avian) antibody
      • V jun sarcoma virus 17 oncogene homolog antibody
      • V-jun avian sarcoma virus 17 oncogene homolog antibody
      • vJun Avian Sarcoma Virus 17 Oncogene Homolog antibody
      see all

    Images

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human lung carcinoma tissue sections labeling c-Jun with purified ab32137 at 1:100 dilution (1.2 µg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
    • Immunocytochemistry/ Immunofluorescence - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Immunocytochemistry/ Immunofluorescence - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Immunocytochemistry/ Immunofluorescence analysis of NIH/3T3 (Mouse embryonic fibroblast) cells labeling c-Jun with purified ab32137 at 1:50 dilution (2.4 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1:200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1:1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
    • Western blot - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Western blot - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Anti-c-Jun antibody [E254] - ChIP Grade (ab32137) at 1/5000 dilution + PC-12(Rat adrenal gland pheochromocytoma) whole cell lysate at 20 µg

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

      Predicted band size: 36 kDa
      Observed band size: 43 kDa
      why is the actual band size different from the predicted?

    • Immunocytochemistry/ Immunofluorescence - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Immunocytochemistry/ Immunofluorescence - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)

      Unpurified ab32137 staining c-Jun in HeLa cells treated with curcumin (diferuloylmethane) (ab120618), by ICC/IF. Decrease in c-Jun expression correlates with increased concentration of curcumin (diferuloylmethane) as described in literature.
      The cells were incubated at 37°C for 4h in media containing different concentrations of ab120618 (curcumin (diferuloylmethane)) in DMSO, fixed with 4% formaldehyde for 10 minutes at room temperature and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab32137 (1/100 dilution) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 goat anti-rabbit polyclonal antibody (ab96899) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.

    • Western blot - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Western blot - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      All lanes : Anti-c-Jun antibody [E254] - ChIP Grade (ab32137) at 1/200 dilution

      Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates prepared in RIPA lysis method
      Lane 2 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates prepared in 1% SDS Hot lysis method.
      Lane 3 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysates prepared in RIPA lysis method
      Lane 4 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysates prepared in 1% SDS Hot lysis method 15ug.

      Lysates/proteins at 15 µg per lane.

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

      Predicted band size: 36 kDa
      Observed band size: 39 kDa why is the actual band size different from the predicted?


      Exposure time: 3 minutes


      Blocking and diluting buffer: 5% NFDM/TBST

      For Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).

    • Immunoprecipitation - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Immunoprecipitation - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)

      ab32137 (purified) at 1/60 dilution (20 µg/mL) immunoprecipitating c-Jun in HEK-293 whole cell lysate.
      Lane 2 (+): HEK-293(Human embronic kidney epithelial cell) whole cell lysate 10 µg
      Lane 3 (-): ab32137 & HEK-293 whole cell lysateRabbit monoclonal IgG (ab172730) instead of ab32137 in HEK-293 whole cell lysate
      For western blotting, ab32137 at 1/500 dilution (2.29 µg/mL) and
      VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1/1000 dilution.
      Blocking and diluting buffer: 5% NFDM /TBST .

    • ChIP - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      ChIP - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Chromatin was prepared from PC-12 (starve overnight) + NGF ( 50 ng/ml 2h) cells according to the Abcam X-ChIP protocol. Cells were fixed with 1% formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 5µg of ab32137 (blue), and 20µl of protein A/G sepharose beads slurry (10µl of sepharose A beads + 10µl of sepharose G beads). 5µg of rabbit normal IgG was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).
    • Immunoprecipitation - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Immunoprecipitation - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      c-Jun was immunoprecipitated using 0.5mg NIH3T3 whole cell extract, 5µg of Rabbit polyclonal to c-Jun 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, NIH3T3 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 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab32137.
      Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).
      Band: 45kDa; c-Jun
    • Immunocytochemistry/ Immunofluorescence - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Immunocytochemistry/ Immunofluorescence - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)

      ICC/IF image of unpurified ab32137 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab32137, 1/200 dilution) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

    • Other - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Other - Anti-c-Jun antibody [E254] - ChIP Grade (ab32137)
      Equilibrium disassociation constant (KD)
      Learn more about KD

      Click here to learn more about KD

    Protocols

    References

    This product has been referenced in:

    • Lakkappa N  et al. Soluble epoxide hydrolase inhibitor, APAU, protects dopaminergic neurons against rotenone induced neurotoxicity: Implications for Parkinson's disease. Neurotoxicology 70:135-145 (2019). Read more (PubMed: 30472438) »
    • Li Y  et al. Overexpression of Opa interacting protein 5 increases the progression of liver cancer via BMPR2/JUN/CHEK1/RAC1 dysregulation. Oncol Rep 41:2075-2088 (2019). Read more (PubMed: 30816485) »
    See all 50 Publications for this product

    Publishing research using ab32137? Please let us know so that we can cite the reference in this datasheet.

    Customer reviews and Q&As

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    1-3 of 3 Abreviews or Q&A

    Western blot abreview for Anti-c-Jun antibody [E254]

     Excellent
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (IM9 cells)
    Gel Running Conditions
    Reduced Denaturing (4-20% Tris Glycin Gel, semi dry transfer on PVDF membrane)
    Loading amount
    25 µg
    Specification
    IM9 cells
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
    Read More

    Abcam user community

    Verified customer

    Submitted Mar 26 2019

    Immunocytochemistry/ Immunofluorescence abreview for Anti-c-Jun antibody [E254] - ChIP Grade

     Excellent
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (AML-193)
    Permeabilization
    Yes - 0.1% Tritonx-100 10min
    Specification
    AML-193
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 20°C
    Fixative
    Formaldehyde
    Read More

    Abcam user community

    Verified customer

    Submitted Sep 12 2017

    Question

    about antibodies to be used in ChIP

    Read More
    Answer

    Thank you very much for your call this morning.
    As discussed, I would be happy to provide you with our testing discount to test one of our anti cFos or cJun antibodies in ChIP.
    This morning we discussed the antibodies anti cJun ab32137, anti cFos ab7963 and anti cFos ab129361.
    https://www.abcam.com/index.html?datasheet=32137 https://www.abcam.com/index.html?datasheet=7963
    https://www.abcam.com/index.html?datasheet=129361
    The testing discount I can offer for each of these antiboides is a discount off a future purchase if you buy one of these antibodies now, test it in ChIP 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 which antibodies you would like to test in ChIP. I will then send a discount code for each of the antibodies. This code must be issued before purchasing the antibodies, so please wait for my reply before ordering.
    2. Purchase the antibodies either by phone, fax, or online (www.abcam.com).
    3. Test it in ChIP.
    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.
    We are always pleased to obtain feedback about our products and any information is greatly appreciated! Even if an antibody turns out to be unsuitable for ChIP, 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.

    Read More

    Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
    For licensing inquiries, please contact partnerships@abcam.com