Anti-p21 antibody [EPR18021] Recombinant (ab188224)

Product name

Anti-p21 antibody [EPR18021]
See all p21 primary antibodies
Description

Rabbit monoclonal [EPR18021] to p21
Host species

Rabbit
Specificity

Expression levels of the target protein vary between different tissue/cell lines and in some cases induction may be required before a signal is observed.
Tested applications

Suitable for: WB, IHC-P, Flow Cyt, IP, ICC/IFmore details
Species reactivity

Reacts with: Mouse
Immunogen

Recombinant full length protein within Mouse p21 aa 1 to the C-terminus. The exact sequence is proprietary.
Database link: P39689

Run BLAST with Run BLAST with
Positive control
- WB: RAW 264.7 whole cell lysate; NIH/3T3, untreated and treated with 1µM staurosporine for 2hrs, whole cell lysates. IHC-P: Mouse testis and lung tissues. ICC/IF: RAW 264.7 and NIH/3T3 cells. Flow Cyt: RAW 264.7 cells. IP: NIN/3T3 whole cell lysate.
General notes

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

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
Storage buffer

Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 0.05% BSA, 40% Glycerol
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR18021
Isotype

IgG
Research areas

Alternative Versions
- Anti-p21 antibody [EPR18021] - BSA and Azide free (ab232512)
Compatible Secondaries
Related Products
- VeriBlot for IP Detection Reagent (HRP) (ab131366)
- Antigen Retrieval Buffer (100X Tris-EDTA Buffer, pH 9.0) (ab93684)

Our Abpromise guarantee covers the use of ab188224 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. Detects a band of approximately 18 kDa (predicted molecular weight: 18 kDa).
IHC-P		1/1000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Flow Cyt		1/50. ab172730-Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
IP		1/30.
ICC/IF		1/500.

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.
Target information above from: UniProt accession P38936 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 12575 Mouse
- SwissProt: P39689 Mouse
- Unigene: 195663 Mouse
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

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR18021] (ab188224)

Immunohistochemical analysis of paraffin-embedded mouse testis tissue labeling p21 with ab188224 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Nuclear staining on mouse testis is observed (PMID: 9170103).

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
Immunocytochemistry/ Immunofluorescence - Anti-p21 antibody [EPR18021] (ab188224)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) cells labeling p21 with ab188224 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on RAW264.7 cells.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.
Western blot - Anti-p21 antibody [EPR18021] (ab188224)

Anti-p21 antibody [EPR18021] (ab188224) at 1/1000 dilution + RAW264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate at 10 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 18 kDa
Observed band size: 18 kDa

Exposure time: 8 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-p21 antibody [EPR18021] (ab188224)

All lanes : Anti-p21 antibody [EPR18021] (ab188224) at 1/1000 dilution

Lane 1 : Untreated NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate
Lane 2 : NIH/3T3 (Mouse embryonic fibroblast cell line) treated with 1µM staurosporine for 2hrs, whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 18 kDa
Observed band size: 18 kDa

Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.

The expression level of p21 protein can be induced using staurosporine (protein kinase C inhibitor) (PMID:7677742).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR18021] (ab188224)

Immunohistochemical analysis of paraffin-embedded mouse lung tissue labeling p21 with ab188224 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Sporadic nuclear staining on mouse lung is observed (PMID: 25333671).

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.
Immunocytochemistry/ Immunofluorescence - Anti-p21 antibody [EPR18021] (ab188224)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling p21 with ab188224 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on NIH/3T3 cells.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.
Flow Cytometry - Anti-p21 antibody [EPR18021] (ab188224)

Flow cytometric analysis of 4% paraformaldehyde-fixed RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) cells labeling p21 with ab188224 at 1/50 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody.
Immunoprecipitation - Anti-p21 antibody [EPR18021] (ab188224)

p21 was immunoprecipitated from 0.35 mg of NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate with ab188224 at 1/30 dilution.

Western blot was performed from the immunoprecipitate using ab188224 at 1/500 dilution.

VeriBlot for IP secondary antibody (HRP) (ab131366), was used as secondary antibody at 1/1000 dilution.

Lane 1: NIH/3T3 whole cell lysate 10 µg (Input).

Lane 2: ab188224 IP in NIH/3T3 whole cell lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab188224 in NIH/3T3 whole cell lysate.

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 10 seconds.

Flow cytometry protocols
Immunoprecipitation protocols
Immunohistochemistry protocols
Immunocytochemistry & immunofluorescence protocols
Western blot protocols

This product has been referenced in:

Jin Y et al. Double deletion of tetraspanins CD9 and CD81 in mice leads to a syndrome resembling accelerated aging. Sci Rep 8:5145 (2018). WB . Read more (PubMed: 29572511) »
Jurisevic M et al. The organic ester O,O'-diethyl-(S,S)-ethylenediamine-N,N'-di-2-(3-cyclohexyl)propanoate dihydrochloride attenuates murine breast cancer growth and metastasis. Oncotarget 9:28195-28212 (2018). Read more (PubMed: 29963272) »

See all 4 Publications for this product

Publishing research using ab188224? Please let us know so that we can cite the reference in this datasheet.

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1-2 of 2 Abreviews

Application

Immunocytochemistry/ Immunofluorescence

Sample

Human Cell (HeLa)

Permeabilization

Yes - 0.5% Triton X-100

Specification

HeLa

Fixative

Paraformaldehyde

Dr. Kirk Mcmanus

Verified customer

Submitted Aug 10 2018

Abcam guarantees this product to work in the species/application used in this Abreview.

Application

Western blot

Sample

Mouse Cell lysate - whole cell (ameloblast)

Gel Running Conditions

Reduced Denaturing (4-15%)

Loading amount

10 µg

Specification

ameloblast

Blocking step

Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Sep 23 2017

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

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Anti-p21 antibody [EPR18021] (ab188224)

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