Overview

  • Product name

    Anti-p21 (phospho T145) antibody
    See all p21 primary antibodies
  • Description

    Rabbit polyclonal to p21 (phospho T145)
  • Host species

    Rabbit
  • Tested applications

    Suitable for: ICC/IF, WB, IHC-P, ELISAmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    synthesized phosphopeptide derived from human p21Cip1 around the phosphorylation site of threonine 145 (R-Q-TP-S-M)

  • Positive control

    • Human breast carcinoma tissue and EGF treated HeLa cell extracts
  • General notes


    p21Cip1 (phospho-Thr145) antibody detects endogenous levels of p21Cip1 only when phosphorylated at threonine 145.

Properties

Applications

Our Abpromise guarantee covers the use of ab47300 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
ICC/IF Use a concentration of 1 µg/ml.
WB 1/500 - 1/1000. Detects a band of approximately 32 kDa (predicted molecular weight: 18 kDa).
IHC-P Use at an assay dependent concentration.
ELISA 1/10000.

Target

  • Function

    May be the important intermediate by which p53/TP53 mediates its role as an inhibitor of cellular proliferation in response to DNA damage. Binds to and inhibits cyclin-dependent kinase activity, preventing phosphorylation of critical cyclin-dependent kinase substrates and blocking cell cycle progression. Functions in the nuclear localization and assembly of cyclin D-CDK4 complex and promotes its kinase activity towards RB1. At higher stoichiometric ratios, inhibits the kinase activity of the cyclin D-CDK4 complex.
  • Tissue specificity

    Expressed in all adult human tissues, with 5-fold lower levels observed in the brain.
  • Sequence similarities

    Belongs to the CDI family.
  • Domain

    The PIP-box K+4 motif mediates both the interaction with PCNA and the recuitment of the DCX(DTL) complex: while the PIP-box interacts with PCNA, the presence of the K+4 submotif, recruits the DCX(DTL) complex, leading to its ubiquitination.
    The C-terminal is required for nuclear localization of the cyclin D-CDK4 complex.
  • Post-translational
    modifications

    Phosphorylation of Thr-145 by Akt or of Ser-146 by PKC impairs binding to PCNA. Phosphorylation at Ser-114 by GSK3-beta enhances ubiquitination by the DCX(DTL) complex.
    Ubiquitinated by MKRN1; leading to polyubiquitination and 26S proteasome-dependent degradation. Ubiquitinated by the DCX(DTL) complex, also named CRL4(CDT2) complex, leading to its degradation during S phase or following UV irradiation. Ubiquitination by the DCX(DTL) complex is essential to control replication licensing and is PCNA-dependent: interacts with PCNA via its PIP-box, while the presence of the containing the 'K+4' motif in the PIP box, recruit the DCX(DTL) complex, leading to its degradation.
  • Cellular localization

    Cytoplasm. Nucleus.
  • Information by UniProt
  • Database links

  • Alternative names

    • CAP20 antibody
    • CDK-interacting protein 1 antibody
    • CDKI antibody
    • CDKN1 antibody
    • Cdkn1a antibody
    • CDN1A_HUMAN antibody
    • CIP1 antibody
    • Cyclin Dependent Kinase Inhibitor 1A antibody
    • Cyclin-dependent kinase inhibitor 1 antibody
    • Cyclin-dependent kinase inhibitor 1A (P21) antibody
    • Cyclin-dependent kinase inhibitor 1A (p21, Cip1) antibody
    • DNA Synthesis Inhibitor antibody
    • MDA-6 antibody
    • MDA6 antibody
    • Melanoma differentiation-associated protein 6 antibody
    • Melanoma differentiation-associated protein antibody
    • p21 antibody
    • P21 protein antibody
    • p21CIP1 antibody
    • p21Cip1/Waf1 antibody
    • p21WAF antibody
    • PIC1 antibody
    • SDI1 antibody
    • SLC12A9 antibody
    • WAF1 antibody
    • Wild type p53 activated fragment 1 (WAF1) antibody
    • Wild type p53 activated fragment 1 antibody
    • Wildtype p53-activated fragment 1 antibody
    see all

Images

  • All lanes : Anti-p21 (phospho T145) antibody (ab47300)

    Lane 1 : EGF treated HeLa cells
    Lane 2 : HeLa cells

    Predicted band size: 18 kDa

  • ab47300 staining human breast carcinoma tissue by IHC-P (left hand panel). The right hand panel shows staining in the presence of phospho-peptide.
  • ICC/IF image of ab47300 stained HeLa cells. The cells were 4% PFA 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 (ab47300, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 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.

References

This product has been referenced in:

See all 5 Publications for this product

Customer reviews and Q&As

1-3 of 3 Abreviews or Q&A

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Dog Tissue sections (Skin)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Antigen retrieval (Dako)
Permeabilization
No
Specification
Skin
Fixative
Formaldehyde

Abcam user community

Verified customer

Submitted Nov 06 2017

Answer

Thank you for contacting Abcam. I have looked at the two antibodies you are interested in, ab39409 and ab47300 and since neither has been tested in mouse, we cannot gurantee that they will work for your particular requirments. However, what I can offer is our testing discount program. I have placed the details of the program below: As neither ab39409 or 47300 have not been tested in mouse tissue, I can offer a discount off a future purchase if you buy either ab39409 or 47300 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 ab39409 or 47300 in mouseI will then send a discount code. This code must be issued before purchasing ab(XXX) so please wait for my reply before ordering. 2. Purchase ab39409 or 47300 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. We are always pleased to obtain feedback about our products and any information is greatly appreciated! Even if ab39409 or 47300 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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Answer

Thank you for your enquiry. The reason why the detected and expected bands are different is maybe due to modifications like the following: 1) post-translational modification - e.g. phosphorylation, glycosylation etc which increases the size of the protein 2) post-translation cleavage - e.g. many proteins are synthesized as pro-proteins, and then cleaved to give the active form, e.g. pro-caspases 3) splice variants - alternative splicing may create different sized proteins from the same gene 4) relative charge - the composition of amino acids (charged vs non-charged) multimers - e.g. dimerisation of a protein. This is usually prevented in reducing conditions, although strong interactions can result in the appearance of higher bands I am afraid that we do not know the exact cause of the difference in band size. The image that we have was the outcome our laboratory obtained during their testing. I am sorry I can't be of more assistance in this occasion but should you require any other information, please do not hesitate to contact me.

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