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    p21-antibody-epr362-bsa-and-azide-free-ab218311.pdf

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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

  • Datasheet
  • Certificate of Compliance
Submit a review Submit a question References (23)

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Western blot - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
  • Immunocytochemistry/ Immunofluorescence - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
  • Immunocytochemistry/ Immunofluorescence - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
  • Immunoprecipitation - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
  • Flow Cytometry (Intracellular) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
  • Western blot - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
  • Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR362] to p21 - BSA and Azide free
  • Suitable for: Flow Cyt (Intra), ICC/IF, IP, WB, IHC-P
  • Knockout validated
  • Reacts with: Human

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Overview

  • Product name

    Anti-p21 antibody [EPR362] - BSA and Azide free
    See all p21 primary antibodies
  • Description

    Rabbit monoclonal [EPR362] to p21 - BSA and Azide free
  • 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: Flow Cyt (Intra), ICC/IF, IP, WB, IHC-Pmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: MCF7, HeLa, HEK293, HUVEC, LnCaP, U87 MG or 293T cell lysates. IHC-P: Human cervical carcinoma or papillary carcinoma of thyroid gland tissues. ICC/IF: MCF-7 cells. Flow Cyt (intra): HeLa cells. IP: HEK293 cell lysate.
  • General notes

    ab218311 is the carrier-free version of ab109520.

    Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.

    This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

    Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.

    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.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C. Do Not Freeze.
  • Storage buffer

    pH: 7.20
    Constituent: PBS
  • Carrier free

    Yes
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR362
  • Isotype

    IgG
  • Research areas

    • Cell Biology
    • Cell Cycle
    • Cell Cycle Inhibitors
    • Cip / Kip
    • Epigenetics and Nuclear Signaling
    • DNA / RNA
    • DNA Damage & Repair
    • DNA Damage Response
    • p53
    • Epigenetics and Nuclear Signaling
    • Cell cycle
    • Cell Cycle Inhibitors
    • Cip/Kip
    • Cancer
    • Cell cycle
    • Cell cycle inhibitors
    • Cip/kip
    • Cancer
    • Oncoproteins/suppressors
    • Tumor suppressors
    • p53 pathway
    • Kits/ Lysates/ Other
    • Kits
    • ELISA Kits
    • ELISA Kits
    • Cancer proteins ELISA kits
    • Signal Transduction
    • Antibodies
    • p21

Associated products

  • Alternative Versions

    • Anti-p21 antibody [EPR362] (ab109520)
    • Alexa Fluor® 488 Anti-p21 antibody [EPR362] (ab282187)
  • Compatible Secondaries

    • VeriBlot for IP Detection Reagent (HRP) (ab131366)
    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab97051)
  • Conjugation kits

    • PE / R-Phycoerythrin Conjugation Kit - Lightning-Link® (ab102918)
    • APC Conjugation Kit - Lightning-Link® (ab201807)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Positive Controls

    • HeLa whole cell lysate (ab150035)
    • MCF7 whole cell lysate (ab29537)
    • HeLa whole cell lysate (ab29545)
    • HEK-293 whole cell lysate (ab7902)
  • Recombinant Protein

    • Recombinant Human p21 protein (ab56278)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab218311 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
Flow Cyt (Intra)
Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

ICC/IF
Use at an assay dependent concentration.

 

IP
Use at an assay dependent concentration.
WB
Use at an assay dependent concentration. Predicted molecular weight: 21 kDa.
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

Notes
Flow Cyt (Intra)
Use at an assay dependent concentration.

ab199376 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

ICC/IF
Use at an assay dependent concentration.

 

IP
Use at an assay dependent concentration.
WB
Use at an assay dependent concentration. Predicted molecular weight: 21 kDa.
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

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.
  • 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: 1026 Human
    • Omim: 116899 Human
    • SwissProt: P38936 Human
    • Unigene: 370771 Human
    • 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

    • Western blot - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Western blot - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      All lanes : Anti-p21 antibody [EPR362] (ab109520) at 1/1000 dilution

      Lane 1 : wild-type HeLa Vehicle Control Fluvastatin (20 uM, 24 h) cell lysate
      Lane 2 : wild-type HeLa Treated Fluvastatin (50 uM, 24 h) cell lysate
      Lane 3 : CDKN1A knockout HeLa Vehicle Control Fluvastatin (20 uM, 24 h) cell lysate
      Lane 4 : CDKN1A knockout HeLa Treated Fluvastatin (50 uM, 24 h) cell lysate

      Performed under reducing conditions.

      Predicted band size: 21 kDa
      Observed band size: 21 kDa



      False colour image of Western blot: Anti-p21 antibody [EPR362] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab109520 was shown to bind specifically to p21. A band was observed at 21 kDa in wild-type HeLa cell lysates with no signal observed at this size in CDKN1A knockout cell line ab255349 (knockout cell lysate ab263812). To generate this image, wild-type and CDKN1A knockout y cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged.Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

    • Immunocytochemistry/ Immunofluorescence - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Immunocytochemistry/ Immunofluorescence - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

      Cell line MCF7 (Human breast adenocarcinoma cell line), Target AbID ab109520  anti-p21 ab150077 AlexaFluor®488 Goat anti-Rabbit secondary. Counterstain AbID ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594). Fixative 4% Paraformaldehyde, Permeabilisation 0.1% tritonX-100, Nuclear counter stain DAPI. Comments Confocal image showing nuclear staining on MCF7 cell line. Target 1oAb dilution 1:500 2 μg/ml, Target 2ndry Ab dilution 1:1000 2 μg/ml, Counterstain Ab dilution 1:200 2.5 μg/ml.

       

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109520).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human thyroid carcinoma tissue labelling p21 with purified ab109520 at 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109520).

    • Immunocytochemistry/ Immunofluorescence - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Immunocytochemistry/ Immunofluorescence - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

      Immunocytochemistry/Immunofluorescence analysis of MCF7 cells labelling p21 with purified ab109520 at 1/1000. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500) were also used.

      Control 1: primary antibody (1/1000) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/500).

      Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/500).

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109520).

    • Immunoprecipitation - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Immunoprecipitation - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

      ab109520 (purified) at 1/50 immunoprecipitating p21 in HEK293 whole cell lysate.

      Lane 1 (input): HEK293 whole cell lysate (10µg)

      Lane 2 (+): ab109520 + HEK293 whole cell lysate (10µg).

      Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab109520 in HEK293 whole cell lysate.

      For western blotting, ab131366 VeriBlot for IP (HRP) was used for detection at 1/1500 dilution.

      Blocking buffer and concentration: 5% NFDM/TBST.

      Diluting buffer and concentration: 5% NFDM /TBST.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109520).

    • Flow Cytometry (Intracellular) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Flow Cytometry (Intracellular) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

      Overlay histogram showing HeLa cells stained with unpurified ab109520 (red line). The cells were fixed with 80% methanol (5 min) then permeabilized with 0.1% PBS-Tween 20 for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (unpurified ab109520, 1/100) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150081) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (ab172730, 1µg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. 

      Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. 

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109520). 

       

       

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervical carcinoma tissue labelling p21 with unpurified ab109520 at a dilution of 1/100.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109520).

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    • Western blot - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Western blot - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      This data was developed using ab109520, the same antibody clone in a different buffer formulation. Different batches of ab109520 were tested on MCF7 (Human breast adenocarcinoma epithelial cell) lysate at 0.2 µg/ml. 15 µg of lysate was loaded in each lane. Bands observed at 21 kDa.
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human papillary carcinoma of the thyroid gland tissue labelling p21 with unpurified ab109520 at a dilution of 1/100.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109520).

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    • Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)
      Anti-p21 antibody [EPR362] - BSA and Azide free (ab218311)

    Protocols

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

    Click here to view the general protocols

    Datasheets and documents

    • Datasheet download

      Download

    Certificate of Compliance

    To download a Certificate of Compliance, please enter your Lot number below:

    References (23)

    Publishing research using ab218311? Please let us know so that we can cite the reference in this datasheet.

    ab218311 has been referenced in 23 publications.

    • Wang L  et al. LncRNA BCYRN1-induced autophagy enhances asparaginase resistance in extranodal NK/T-cell lymphoma. Theranostics 11:925-940 (2021). PubMed: 33391513
    • Wu J  et al. LINC01152 upregulates MAML2 expression to modulate the progression of glioblastoma multiforme via Notch signaling pathway. Cell Death Dis 12:115 (2021). PubMed: 33483471
    • Shen H  et al. Enhancing the potential of aged human articular chondrocytes for high-quality cartilage regeneration. FASEB J 35:e21410 (2021). PubMed: 33617078
    • Liu W  et al. miR-140 inhibits osteosarcoma progression by impairing USP22-mediated LSD1 stabilization and promoting p21 expression. Mol Ther Nucleic Acids 24:436-448 (2021). PubMed: 33868787
    • Cao D  et al. Phorbol myristate acetate induces cellular senescence in rat microglia in vitro. Int J Mol Med 46:415-426 (2020). PubMed: 32626908
    View all Publications for this product

    Customer reviews and Q&As

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