概述

  • 产品名称
    Anti-CDKN2A/p19ARF抗体
    参阅全部 CDKN2A/p19ARF 一抗
  • 描述
    兔多克隆抗体to CDKN2A/p19ARF
  • 宿主
    Rabbit
  • 特异性
    On Western blot the antibody reveals IVT murine p19ARF and a band of the same size in CTLL2 cells. It detects a specific band in WT mouse embryo fibroblasts which is not present in p19ARF-null MEFs.
  • 经测试应用
    适用于: ICC/IF, ICC, ELISA, IHC-P, WB, IHC-Frmore details
  • 种属反应性
    与反应: Mouse
    不与反应: Rat, Human
  • 免疫原

    Synthetic peptide conjugated to KLH derived from within residues 50 - 150 of Mouse p19ARF.

    参阅Abcam的专有抗源政策 (Peptide available as ab2659.)

  • 阳性对照
    • mouse embryonic fibroblasts lysate
  • 常规说明
    The blocking peptide for this antibody is available (ab2659)

性能

应用

Our Abpromise guarantee covers the use of ab80 in the following tested applications.

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

应用 Ab评论 说明
ICC/IF 1/250.
ICC Use a concentration of 1 µg/ml. Fix with a 1:1 mixture of methanol:acetone at -20C for 10 mins (see image by Dr. David Bertwistle).
ELISA Use at an assay dependent concentration.
IHC-P Use a concentration of 1 µg/ml. PubMed: 11726500
WB 1/1000. Detects a band of approximately 19 kDa (predicted molecular weight: 17 kDa). Multiple bands with this antibody have frequently been reported. We believe that this is due to the different modification states of p19ARF. Nuclear extraction may be necessary in some samples.
IHC-Fr 1/100.

靶标

  • 相关性
    The gene for CDK2NA generates several transcripts/proteins which differ from each other in their first exons. Three of these transcripts are generated by alternative splicing (isoform 1 a.k.a p16INK4A, isoform 2 and isoform 3 a.k.a p12), two of which are known to function as inhibitors of CDK4 kinase. One other transcript that is generated from this gene contains an alternate reading frame (ARF), with the first exon located 20kb upstream of the remainder of the gene(isoform 4 a.k.a. p14ARF, p19ARF, ARF). In spite of the structural and some functional differences, all the proteins encoded by the CDKN2A gene are involved in cell cycle G1 control.
  • 细胞定位
    Nuclear
  • 数据库链接
  • 别名
    • ARF antibody
    • CDKN2A antibody
    • Cyclin dependent kinase inhibitor 2A antibody
    • INK4 antibody
    • MTS1 antibody
    • P19 antibody
    see all

图片

  • All lanes : Anti-CDKN2A/p19ARF antibody (ab80) at 1 µg/ml

    Lanes 1-2 : 30 ug of mouse embryonic fibroblast total cell extract.
    Lane 3 : 30 ug of mouse embryonic fibroblast total cell extract, with the p19ARF gene knocked out as a negative control.

    Performed under reducing conditions.

    Predicted band size: 17 kDa
    Observed band size: 19 kDa
    why is the actual band size different from the predicted?



    Western blot of primary Mouse Embryo Fibroplasts (MEFs) using ab80 at 1 µg/ml.  (30 µg of protein from total cell extracts per lane.)

  • ab80 antibody was tested on NIH 3T3 cells (which have a deletion of the p19ARF gene) as a negative control (-) and NIH 3T3 cells engineered to express HA-tagged p19ARF as a positive control (+).

    Cells grown on coverslips were fixed with a 1:1 mixture of methanol:acetone at -20C for 10 mins. After drying, rehydrating and blocking, the co

  • ab80 at 1/300 staining mouse kidney (glomerulus) tissue sections by IHC-P. The tissue was formaldehyde fixed and a heat mediated antigen retrieval step in citrate buffer was performed. The tissue was then blocked and incubated with the antibody for 45 minutes. An HRP conjugated goat anti-rabbit antibody was used as the secondary.

    See Abreview

  • ab80 at 1/300 staining mouse brain cells by ICC/IF.

    Paraformaldehyde fixation, Triton X-100 permeabilization, followed by blocking with 3% BSA + 2% NGS for 1 hour at 20°C.

    Incubation with the primary antibody was carried out for 1 hour at 20°C.  The secondary antibody was an Alexa Fluor® conjugated Donkey anti-Rabbit polyclonal, diluted 1/400.

    See Abreview

  • ab80 staining CDKN2A/p19ARF in Mouse pancreatic neoplastic tissue sections by Immunohistochemistry (IHC-Fr - frozen sections). Tissue was fixed with paraformaldehyde and blocked with 5% serum for 1 hour at room temperature. Samples were incubated with primary antibody (1/100 in PBS) for 8 hours at 4°C. An Alexa Fluor®555-conjugated Donkey anti-rabbit IgG polyclonal (1/500) was used as the secondary antibody. Nuclei were stained by DAPI.

    See Abreview

文献

This product has been referenced in:
  • Oldrini B  et al. Somatic genome editing with the RCAS-TVA-CRISPR-Cas9 system for precision tumor modeling. Nat Commun 9:1466 (2018). Read more (PubMed: 29654229) »
  • Francis JC  et al. SOX9 is a driver of aggressive prostate cancer by promoting invasion, cell fate and cytoskeleton alterations and epithelial to mesenchymal transition. Oncotarget 9:7604-7615 (2018). Read more (PubMed: 29484136) »
See all 195 Publications for this product

客户评价及客户问答

1-10 of 37 Abreviews or Q&A

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

Abcam user community

Verified customer

提交于 Aug 20 2018

Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (HF)
Permeabilization
Yes - PBS-TWEEN 0.05%
Specification
HF
Blocking step
Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C
Fixative
Acetone

Abcam user community

Verified customer

提交于 Feb 23 2018

Abreviews
Application
IHC - Wholemount
Sample
Mouse Tissue (inner ear tissue)
Specification
inner ear tissue

Abcam user community

Verified customer

提交于 Jun 11 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - whole cell (mouse whole cell lysate)
Loading amount
30 µg
Specification
mouse whole cell lysate
Gel Running Conditions
Reduced Denaturing (4-20%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Abcam user community

Verified customer

提交于 Apr 05 2013

Answer

Thank you for contacting us. Depending on the gel, samples, and buffer, the bands can appear 10 kD from the "predicted" size on your gel. Since the concentration of your antibody is 0.6 mg/ml and we suggest using 1 ug/ml on your blot, you could try using 1/500 primary overnight to see if any other bands appear. You could also use the blocking peptide ab2659 to see if the band on your blot is specifically blocked.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information. We do guarantee our products to work as specified on the datasheet and can replace, credit or refund the product within 6 months if you are not satisfied with your results.

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

Read More

Answer

Thank you for contacting us.

Based on this image alone it seems like the lower band is your CDKN2A/p19ARF band. It is possible for band size to change slightly due to sample preparation or buffer, however usually the positive control will be a good indicator of where the protein of interest is. One strategy to confirm the band might be to use the blocking peptide (the peptide used as the immunogen) to specifically block the CDKN2A/p19ARF protein. We have this available as ab
2659.


I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

Read More

Answer

Thank you very much for contacting us with your enquiry.

I'm sorry to hear that there were no bands in Western blot using this antibody. I would expect any of the MEF cell lines to work as a positive control, but I suggest using a whole cell lysate (ab14874 is a nuclear fraction) since that is what we have tested with ab80. What kind of samples are you using other than the MEF lysate?

If you'd like to send some more details about your protocol ( how much protein was loaded, dilution of antibody, blocking solution, etc) I'd be happy to take a look and see if we have any suggestions to improve the results.

I look forward to hearing from you. If there's anything else that we can do for you, please let me know and I'll be happy to help.

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Question
Answer

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products. I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement of ab80 from a different lot. To check the status of the order please contact our Customer Service team and reference this number. Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know. I wish you the best of luck with your research.  

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Answer

My colleague from US Customer Service has passed your message to me. Thank you for your recent call regarding ab80. I am very sorry to hear that you are having problems with this antibody. All our customer feedback, including complaints are monitored weekly by our in house technical support team. If a product is at fault the technical support team will consider removing the product from our catalogue in order to avoid future customer inconvenience. We are not aware of any batch-related quality-problem. Could you please check and confirm in what species (sample, cell or tissue types) you have been using this antibody which failed to work? I look forward to hearing from you soon.  

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (mouse pancreatic cancer primary)
Specification
mouse pancreatic cancer primary
Fixative
Acetone
Permeabilization
Yes - PBS + Triton 0.025%
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: RT°C

Abcam user community

Verified customer

提交于 Jul 19 2010

1-10 of 37 Abreviews or Q&A

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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