製品の概要

  • 製品名
    Anti-c-Jun antibody - ChIP Grade
    c-Jun 一次抗体 製品一覧
  • 製品の詳細
    Rabbit polyclonal to c-Jun - ChIP Grade
  • 由来種
    Rabbit
  • 特異性
    Antibody detects endogenous levels of c-Jun protein around Serine 243.
  • アプリケーション
    適用あり: ChIP, ICC/IF, IHC-P, IP, WB, ELISA, Flow Cytmore details
  • 種交差性
    交差種: Mouse, Rat, Human, African green monkey
    交差が予測される動物種: Chicken, Cow, Pig
  • 免疫原

    Synthetic peptide within Human c-Jun aa 210-259. The exact sequence is proprietary.
    Sequence:

    HLPQQMPVQHPRLQALKEEPQTVPEMPGETPPLSPIDMESQERIKAERKR


    Database link: P05412

  • ポジティブ・コントロール
    • ChIP: Human endothelial cells (EA.hy926). IHC-P: Human breast carcinoma tissue. ICC/IF: HepG2 cells. WB: Extracts of HeLa cells. Recombinant Human c-Jun protein. Flow Cytometry: Jurkat cells.

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab31419 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

アプリケーション Abreviews 特記事項
ChIP Use at an assay dependent concentration.
ICC/IF Use a concentration of 1 µg/ml.
IHC-P 1/50 - 1/100.
IP Use at an assay dependent concentration.
WB 1/500 - 1/1000. Predicted molecular weight: 36 kDa.
ELISA 1/20000.
Flow Cyt 1/300.

ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.

ターゲット情報

  • 機能
    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).
  • 配列類似性
    Belongs to the bZIP family. Jun subfamily.
    Contains 1 bZIP (basic-leucine zipper) domain.
  • 翻訳後修飾
    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.
  • 細胞内局在
    Nucleus.
  • Information by UniProt
  • 参照データベース
  • 別名
    • Activator protein 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

画像

  • ChIP analysis using ab31419 binding c-Jun in human endothelial cells (EA.hy926). Cells were cross-linked for 10 minutes with formaldehyde then incubated with undiluted primary antibody for 10 hours at 4°C in 1x ChIP buffer. Protein binding was detected using real-time PCR.
    Positive control: Position 89340150-89340297 in chromosome 11 (has a validated c-Jun site).
    Negative Control: Igr5 intron 3 (contains no c-Jun binding site).

    See Abreview

  • Paraffin-embedded human breast carcinoma tissue stained for c-Jun with ab31419 at a 1/50 dilution in immunohistochemical analysis.

    Left panel: Untreated.

    Right panel: Pre-incubated with synthesized peptide.

  • ICC/IF image of  HepG2 (Human liver hepatocellular carcinoma cell line) cells labeling c-Jun (green) with ab31419 at 1 µg/ml. The cells were fixed in 4% PFA (10 minutes) and then incubated in 1% BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1 hour to permeabilize the cells and block non-specific protein-protein interactions. The cells were then incubated with ab31419 at 1 µg/ml overnight at +4 °C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) ab150077 used at a 1/1000 dilution for 1 hour. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1 hour. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43 µM.

  • All lanes : Anti-c-Jun antibody - ChIP Grade (ab31419) at 1/500 dilution

    Lane 1 : Extracts of Hela (Human epithelial cell line from cervix adenocarcinoma) cells
    Lane 2 : Extracts of Hela (Human epithelial cell line from cervix adenocarcinoma) cells with immunizing peptide

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

  • c-Jun was immunoprecipitated from HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate using ab31419 at a 1/500 dilution.

    Lane 1: Control IgG IP in HEK-293T whole cell lysate.

    Lane 2: ab31419 in HEK-293T whole cell lysate.

    Lane 3: c-Jun in HEK-293T whole cell lysate 500 µg (Input).

    For western blotting an HRP-conjugated swine anti-rabbit polyclonal was used as the secondary antibody.

    See Abreview

  • Overlay histogram showing Jurkat (Human T cell leukemia cell line from peripheral blood) cells stained with ab31419 (blue line) at a 1/300 dilution. The cells were prepared as follows: Spin down, wash in FACS buffer 1x, fix, wash 2x, and stain with primary antibody overnight. Buffer was 0.1% sodium azide with FBS in phosphate buffered solution. Cells were fixed using paraformaldehyde and permeabilized using Triton X-100 and NP40. Cells were gated by isolating cell population from plot of SSC-A / FSA-A. The secondary antibody used was a FITC-conjugated Goat anti-Rabbit polyclonal, diluted 1/100.

    See Abreview

  • Anti-c-Jun antibody - ChIP Grade (ab31419) at 1/500 dilution + Recombinant Human c-Jun protein (ab54318) at 0.01 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 36 kDa


    Exposure time: 30 seconds

参考文献

This product has been referenced in:
  • Yu CY  et al. MicroRNA-125b-5p improves pancreatic ß-cell function through inhibiting JNK signaling pathway by targeting DACT1 in mice with type 2 diabetes mellitus. Life Sci N/A:N/A (2019). Read more (PubMed: 30684546) »
  • Lin B  et al. c-Jun suppresses the expression of WNT inhibitory factor 1 through transcriptional regulation and interaction with DNA methyltransferase 1 in gallbladder cancer. Mol Med Rep 17:8180-8188 (2018). Read more (PubMed: 29693707) »
See all 41 Publications for this product

レビューと Q&A

1-10 of 21 Abreviews or Q&A

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Na-citrate pH6
Sample
Mouse Tissue sections (Liver, P5)
Specification
Liver, P5
Permeabilization
Yes - Triton 0,05%
Fixative
Paraformaldehyde

Abcam user community

Verified customer

投稿 Oct 07 2014

Application
Immunoprecipitation
Immuno-precipitation step
Other - EMSA
Sample
Human Cell lysate - nuclear (renal cancer cells)
Specification
renal cancer cells
Total protein in input
3 µg

Abcam user community

Verified customer

投稿 Mar 14 2014

ChIP

Excellent
Abreviews
Application
ChIP
Detection step
Real-time PCR
Sample
Human Cell lysate - whole cell (EA.hy926 endothelial cells)
Specification
EA.hy926 endothelial cells
Negative control
Igr5 intron 3 contains no c-Jun binding site (Aguilera C et al. Nature. 2011 469: 231-235)
Type
Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
Specification of the cross-linking agent: formaldehyde (final concentrat
Positive control
Position 89340150-89340297 in chromosome 11 has a validated c-Jun site (ENCODE Project Consortium. PLoS Biol. 2011 9:e1001046

Abcam user community

Verified customer

投稿 Jun 05 2013

Western blot

Excellent
Abreviews
Application
Western blot
Loading amount
30 µg
Gel Running Conditions
Reduced Denaturing (12%)
Sample
Human Cell lysate - whole cell (EA.hy926 endothelial cells)
Specification
EA.hy926 endothelial cells
Treatment
c-Jun siRNA for 48h
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

投稿 Jun 05 2013

Question

Hello , Here is the complain of two antibodies .

1) Abcam catalogue no:

Ab31419, Ab47476

2) a) Abcam order number (not required):

1148646


b) Abcam product lot number:

c-Jun> GR50836-9, ATF-2> GR72060-1

3) Description of the problem (high background, wrong band size, more bands, no band etc):

No bands

4) Antibody storage conditions (temperature/reconstitution etc):

-20°C


5) a)Sample (Species):


c-Jun and ATF2 from HeLa


b) Type of sample (Cell extract/Nuclear extract/Purified protein/Recombinant protein etc).:


Whole cell extract

6) a) Sample preparation:

Lysis buffer



RIPA





Protease inhibitors



AEBSF, 2 mM
Aprotinin, 0.3 μM
Bestatin, 130 μM
EDTA, 1 mM
E-64, 14 μM
Leupeptin, 1 μM

PMSF, 20 mM





Phosphatase inhibitors



-





Reducing agent



SDS and β-ME





Boiling for ≥5 min?



5 minutes







b) Amount of protein loaded:







Protein loaded ug/lane or cells/lane



5 μg/lane







7) a) Electrophoresis/Gel conditions:







Reducing or Non Reducing gel



Reducing





Percentage of gel



15%





Volts applied



75





Time applied



3 hours

















b) Transfer or blocking conditions:







Type of membrane



PVDF





Protein transfer verified



Ponceau, β-actin





Blocking agent and concentration



3% BSA





Blocking time



30 minutes





Blocking temperature



25°C

















8) Primary antibody (If more than one was used, describe in “additional notes”):









Concentration or dilution



1/1000





Diluent buffer



1% BSA + 0.05% Tween-20 in 1X PBS





Incubation time



2 hours





Incubation temperature



25°C





Washing: Buffer Used



0.05% Tween-20 in 1X PBS





Number of washes



3







9) Secondary antibody:









Species



Goat





Reacts against



Rabbit





Concentration or dilution



1/10,000





Diluent buffer



1% BSA + 0.05% Tween-20 in 1X PBS





Incubation time



2 hours





Incubation temperature:



25°C





Fluorochrome or enzyme conjugate



HRP





Washing: Buffer Used



0.05% Tween-20 in 1X PBS





Number of washes



3







10) Detection method (ECL, ECL plus, etc.):









ECL





11) Positive and negative controls used:









Positive control



β-actin





Negative control











12) a) How many times you have run this staining?







Twice







b) Do you obtain the same results every time?







No







C) What steps have you altered to try and optimize the use of this antibody?







Reducing blocking, primary and secondary incubation times while attempting the second time. Washing was made slightly rigorous as compared to he first time. Performed a Dot blot to check for performance of secondary antibody and it works fine. Increased amount of loaded protein to 10 μg/lane.







Regards

Read More
Answer

Thank you for your enquiry regarding ab31419 and ab47476; and for taking the time to provide some useful details of the experiments. I am very sorry to hear that your customer is having problems with these two antibodies.
I would like to reassure you that our Abpromise applies to your complaint since you purchased this product within the guarantee period. This means that in the event that a product is not functioning in the applications/species cited on the product data sheet (and the problem has been reported within 6 months of purchase) we will happily offer a credit note/refund to the value of the product purchased.
After reading through the detailed protocol you kindly forwarded to Abcam, I would like to make the following comments/suggestions:
1) The loaded amount of protein seems to be a bit low: 5 μg/lane, normally we would suggest loading at least 25-30 μg/lane protein per lane.
2) Since c-Jun and ATF2 are both mainly present in the nuclei, it would be worth preparing nuclear fraction (rather than whole cell lysate) to enrich the target proteins in the sample if it is possible and apply them for WB.
3) Customer may also need to test different dilutions (1/500 or even 1/250) to make sure that the experiment is carried out under the optimal conditions. This is even more important since only 5 μg instead of 30 μg lysate was loaded.
I hope this will be useful for you. Should you still have any problem with this antibody after following these suggestions, then please do not hesitate to contact our Technical Department again.

Read More

Answer

Thank you for contacting us.

I do wish to aplogize for the delays in getting information about this product to you. I have been informded by the originator of this product that the images are using ductal breast carcinoma however as of this time we do not have any information regarding the percentages of breat carcinoma would be positive.



This product is covered by our Abpromise guarantee. We are happy provide scientific support at any time. If you are using the product in species and applications listed on the datasheet and contact us within six months of purchase, we are also happy to replace or refund the product should we not be able to help you to resolve the issue. More information on our Abpromise may be found at the following link:

https://www.abcam.com/index.html?pageconfig=abpromise

I hope that this information is helpful. Please let me know if you have any questions or there are other ways that Abcam may help you meet your research goals

Free Rabbit monoclonal antibody with any purchase of a primary antibody, while stocks last! Quote “RABMAB-XBSMG” in your next primary antibody order. For more information, visit the following link: https://www.abcam.com/index.html?pageconfig=resource&rid=15447

Read More

Answer

Thank you very much for contacting us.

I just wanted to let you know that I am still working on your inquiry. I have contacted the originator of this product for the information you requested, and I am awaiting a reply.

I can provide some information regarding postive controls for this product. This antibody has been tested successfully in the following tissues/cell lines as shown in on the datasheet and within publicatiobns referenced:

Hela, Jurkat, Mouse small bowel adenoma, U2OS, 3T3, COS7, PC12 , Primary Prostate Epithelia [PMID: 19592505], dorsal mouse skin [PMID: 19762425]


I am very sorry for the delay. I will forward to you the information as soon as I receive it. Thank you for your patience.

Read More
Application
Western blot
Sample
Human Cell lysate - whole cell (HCT116)
Loading amount
30 µg
Specification
HCT116
Gel Running Conditions
Non-reduced Denaturing (12%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

投稿 Nov 07 2012

Answer

Thank you for your reply.

Unfortunately we do not have another C-Jun antibody that is tested in ChIP. Please let me know how you would like to proceed. We do not currently stock a C-Fos antibody that will work in ChIP but if you would like to test an antibody then please let me know. You may be eligible for a testing discount.

I look forward to your reply.

Read More

Question
Answer

Thank you for your reply and with clarifying information.

We have not received any other complaints about this antibody, so I am sorry to hear you have been experiencing problems. It is troubling that it is not working in both ChIP and WB. I would like to offer you a replacement vial of the same or an alternative product, or a refund/credit. Please could you confirm your preference and also provide the following details:

- Date of order/delivery
- Primary antibody dilution

I look forward to your reply.

Read More

1-10 of 21 Abreviews or Q&A

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