Overview

  • Product nameAnti-p53 antibody [PAb 1801]
    See all p53 primary antibodies
  • Description
    Mouse monoclonal [PAb 1801] to p53
  • SpecificityIn our hands, this product has shown a positive signal in western blot at 53 kDa using Human samples only. Some customers have successfully used this product in Mouse and Rat. The following publications suggest that this clone may cross react in ICC with an unknown p-body protein (PMID: 11313875; PMCID: PMC3349707). We therefore are no longer able to guarantee this product for use in mouse and rat.
  • Tested applicationsSuitable for: ChIP, ICC/IF, Flow Cyt, WB, IP, ELISA, IHC-P, RIA, IHC-Frmore details
  • Species reactivity
    Reacts with: Human
  • Immunogen

    Fusion protein corresponding to Human p53 (N terminal).
    Database link: P04637

  • Epitopeaa 46-55 of human p53
  • Positive control
    • This antibody gave a positive signal in the following whole cell lysate: MDA-MB-231. IHC-P: Human pancreas adenocarcinoma FFPE tissue sections, Human colon adenocarcinoma FFPE tissue sections
  • General notes

    This monoclonal recognises both wild-type and mutant forms of human p53 protein.Epitope: aa 46-55 of human p53.

     

    Alternative versions available:

    Anti-p53 antibody (Alexa Fluor® 488) [PAb 1801] (ab193581)

    Anti-p53 antibody (Alexa Fluor® 647) [PAb 1801] (ab193587)

    Anti-p53 antibody (HRP) [PAb 1801] (ab193605)

Properties

Applications

Our Abpromise guarantee covers the use of ab28 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
ChIP Use at an assay dependent concentration.

Use at an assay dependent concentration.

 

ICC/IF Use at an assay dependent concentration.
Flow Cyt Use 1µg for 106 cells.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

WB Use a concentration of 5 µg/ml. Detects a band of approximately 53 kDa (predicted molecular weight: 53 kDa).

We recommend using 3% milk as the blocking agent for Western blot.

IP Use at 10 µg/mg of lysate.
ELISA Use at an assay dependent concentration.
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
RIA Use at an assay dependent concentration.
IHC-Fr 1/100 - 1/250.

Target

  • FunctionActs as a tumor suppressor in many tumor types; induces growth arrest or apoptosis depending on the physiological circumstances and cell type. Involved in cell cycle regulation as a trans-activator that acts to negatively regulate cell division by controlling a set of genes required for this process. One of the activated genes is an inhibitor of cyclin-dependent kinases. Apoptosis induction seems to be mediated either by stimulation of BAX and FAS antigen expression, or by repression of Bcl-2 expression. Implicated in Notch signaling cross-over. Isoform 2 enhances the transactivation activity of isoform 1 from some but not all TP53-inducible promoters. Isoform 4 suppresses transactivation activity and impairs growth suppression mediated by isoform 1. Isoform 7 inhibits isoform 1-mediated apoptosis.
  • Tissue specificityUbiquitous. Isoforms are expressed in a wide range of normal tissues but in a tissue-dependent manner. Isoform 2 is expressed in most normal tissues but is not detected in brain, lung, prostate, muscle, fetal brain, spinal cord and fetal liver. Isoform 3 is expressed in most normal tissues but is not detected in lung, spleen, testis, fetal brain, spinal cord and fetal liver. Isoform 7 is expressed in most normal tissues but is not detected in prostate, uterus, skeletal muscle and breast. Isoform 8 is detected only in colon, bone marrow, testis, fetal brain and intestine. Isoform 9 is expressed in most normal tissues but is not detected in brain, heart, lung, fetal liver, salivary gland, breast or intestine.
  • Involvement in diseaseNote=TP53 is found in increased amounts in a wide variety of transformed cells. TP53 is frequently mutated or inactivated in about 60% of cancers. TP53 defects are found in Barrett metaplasia a condition in which the normally stratified squamous epithelium of the lower esophagus is replaced by a metaplastic columnar epithelium. The condition develops as a complication in approximately 10% of patients with chronic gastroesophageal reflux disease and predisposes to the development of esophageal adenocarcinoma.
    Defects in TP53 are a cause of esophageal cancer (ESCR) [MIM:133239].
    Defects in TP53 are a cause of Li-Fraumeni syndrome (LFS) [MIM:151623]. LFS is an autosomal dominant familial cancer syndrome that in its classic form is defined by the existence of a proband affected by a sarcoma before 45 years with a first degree relative affected by any tumor before 45 years and another first degree relative with any tumor before 45 years or a sarcoma at any age. Other clinical definitions for LFS have been proposed (PubMed:8118819 and PubMed:8718514) and called Li-Fraumeni like syndrome (LFL). In these families affected relatives develop a diverse set of malignancies at unusually early ages. Four types of cancers account for 80% of tumors occurring in TP53 germline mutation carriers: breast cancers, soft tissue and bone sarcomas, brain tumors (astrocytomas) and adrenocortical carcinomas. Less frequent tumors include choroid plexus carcinoma or papilloma before the age of 15, rhabdomyosarcoma before the age of 5, leukemia, Wilms tumor, malignant phyllodes tumor, colorectal and gastric cancers.
    Defects in TP53 are involved in head and neck squamous cell carcinomas (HNSCC) [MIM:275355]; also known as squamous cell carcinoma of the head and neck.
    Defects in TP53 are a cause of lung cancer (LNCR) [MIM:211980].
    Defects in TP53 are a cause of choroid plexus papilloma (CPLPA) [MIM:260500]. Choroid plexus papilloma is a slow-growing benign tumor of the choroid plexus that often invades the leptomeninges. In children it is usually in a lateral ventricle but in adults it is more often in the fourth ventricle. Hydrocephalus is common, either from obstruction or from tumor secretion of cerebrospinal fluid. If it undergoes malignant transformation it is called a choroid plexus carcinoma. Primary choroid plexus tumors are rare and usually occur in early childhood.
    Defects in TP53 are a cause of adrenocortical carcinoma (ADCC) [MIM:202300]. ADCC is a rare childhood tumor of the adrenal cortex. It occurs with increased frequency in patients with the Beckwith-Wiedemann syndrome and is a component tumor in Li-Fraumeni syndrome.
  • Sequence similaritiesBelongs to the p53 family.
  • DomainThe nuclear export signal acts as a transcriptional repression domain. The TADI and TADII motifs (residues 17 to 25 and 48 to 56) correspond both to 9aaTAD motifs which are transactivation domains present in a large number of yeast and animal transcription factors.
  • Post-translational
    modifications
    Acetylated. Acetylation of Lys-382 by CREBBP enhances transcriptional activity. Deacetylation of Lys-382 by SIRT1 impairs its ability to induce proapoptotic program and modulate cell senescence.
    Phosphorylation on Ser residues mediates transcriptional activation. Phosphorylated by HIPK1 (By similarity). Phosphorylation at Ser-9 by HIPK4 increases repression activity on BIRC5 promoter. Phosphorylated on Thr-18 by VRK1. Phosphorylated on Ser-20 by CHEK2 in response to DNA damage, which prevents ubiquitination by MDM2. Phosphorylated on Thr-55 by TAF1, which promotes MDM2-mediated degradation. Phosphorylated on Ser-46 by HIPK2 upon UV irradiation. Phosphorylation on Ser-46 is required for acetylation by CREBBP. Phosphorylated on Ser-392 following UV but not gamma irradiation. Phosphorylated upon DNA damage, probably by ATM or ATR. Phosphorylated on Ser-15 upon ultraviolet irradiation; which is enhanced by interaction with BANP.
    Dephosphorylated by PP2A-PPP2R5C holoenzyme at Thr-55. SV40 small T antigen inhibits the dephosphorylation by the AC form of PP2A.
    May be O-glycosylated in the C-terminal basic region. Studied in EB-1 cell line.
    Ubiquitinated by MDM2 and SYVN1, which leads to proteasomal degradation. Ubiquitinated by RFWD3, which works in cooperation with MDM2 and may catalyze the formation of short polyubiquitin chains on p53/TP53 that are not targeted to the proteasome. Ubiquitinated by MKRN1 at Lys-291 and Lys-292, which leads to proteasomal degradation. Deubiquitinated by USP10, leading to its stabilization. Ubiquitinated by TRIM24, which leads to proteasomal degradation. Ubiquitination by TOPORS induces degradation. Deubiquitination by USP7, leading to stabilization. Isoform 4 is monoubiquitinated in an MDM2-independent manner.
    Monomethylated at Lys-372 by SETD7, leading to stabilization and increased transcriptional activation. Monomethylated at Lys-370 by SMYD2, leading to decreased DNA-binding activity and subsequent transcriptional regulation activity. Lys-372 monomethylation prevents interaction with SMYD2 and subsequent monomethylation at Lys-370. Dimethylated at Lys-373 by EHMT1 and EHMT2. Monomethylated at Lys-382 by SETD8, promoting interaction with L3MBTL1 and leading to repress transcriptional activity. Demethylation of dimethylated Lys-370 by KDM1A prevents interaction with TP53BP1 and represses TP53-mediated transcriptional activation.
    Sumoylated by SUMO1.
  • Cellular localizationCytoplasm; Cytoplasm. Nucleus. Nucleus > PML body. Endoplasmic reticulum. Interaction with BANP promotes nuclear localization. Recruited into PML bodies together with CHEK2; Nucleus. Cytoplasm. Localized in both nucleus and cytoplasm in most cells. In some cells, forms foci in the nucleus that are different from nucleoli; Nucleus. Cytoplasm. Localized in the nucleus in most cells but found in the cytoplasm in some cells; Nucleus. Cytoplasm. Localized mainly in the nucleus with minor staining in the cytoplasm; Nucleus. Cytoplasm. Predominantly nuclear but localizes to the cytoplasm when expressed with isoform 4 and Nucleus. Cytoplasm. Predominantly nuclear but translocates to the cytoplasm following cell stress.
  • Information by UniProt
  • Database links
  • Alternative names
    • Antigen NY-CO-13 antibody
    • BCC7 antibody
    • Cellular tumor antigen p53 antibody
    • FLJ92943 antibody
    • LFS1 antibody
    • Mutant tumor protein 53 antibody
    • p53 antibody
    • p53 tumor suppressor antibody
    • P53_HUMAN antibody
    • Phosphoprotein p53 antibody
    • Tp53 antibody
    • Transformation related protein 53 antibody
    • TRP53 antibody
    • Tumor protein 53 antibody
    • Tumor protein p53 antibody
    • Tumor suppressor p53 antibody
    see all

Anti-p53 antibody [PAb 1801] images

  • All lanes : Anti-p53 antibody [PAb 1801] (ab28) at 5 µg/ml

    Lane 1 : Wild-type HCT116 cell lysate at 30 µg
    Lane 2 : Wild-type HCT116 + irinotecan (10 µM, 24 hours) cell lysate at 30 µg
    Lane 3 : p53 knockout HCT116 cell lysate at 30 µg
    Lane 4 : p53 knockout HCT116 + irinotecan (10 µM, 24 hours) cell lysate at 30 µg
    Lane 5 : A431 cell lysate (positive control) at 20 µg
    Lane 6 : Saos-2 cell lysate (negative control) at 20 µg
    Lane 7 : MEF cell lysate at 20 µg
    Lane 8 : Wild-type HAP1 cell lysate at 20 µg
    Lane 9 : p53 knockout HAP1 cell lysate at 20 µg


    Performed under reducing conditions.

    Predicted band size : 53 kDa

    Lanes 1-9: Merged (red and green) signal.
    Ab28 was shown to specifically react with p53, when p53 knockout samples were used. Wild-type and p53 knockout samples, positive and negative controls were subjected to SDS-PAGE. Ab28 and ab181602(loading control to GAPDH) were diluted 5 μg/mL and 1/10,000 respectively and incubated overnight at 4°C. Blots were developed with goat anti-rabbit IgG (H + L) and goat anti-mouse IgG (H + L) secondary antibodies at 1/10,000 dilution for 1 h at room temperature before imaging.

  • IHC image of ab28 staining p53 in human colon adenocarcinoma formalin-fixed paraffin-embedded tissue sections*, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab28, 1/500 dilution, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. High magnification of the tumor region - T (lower right panel) and adjacent normal crypts - N (lower left panel) are shown.
    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

  • IHC image of p53 staining in human pancreas adenocarcinoma formalin fixed paraffin embedded tissue section, performed on a Leica Bond system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab28, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • ab28 staining p53 in HeLa cells treated with genipin (ab141049), by ICC/IF. Increase of p53 expression correlates with increased concentration of genipin, as described in literature.
    The cells were incubated at 37°C for 6h in media containing different concentrations of ab141049 (genipin) in DMSO, fixed with 4% formaldehyde for 10 minutes at room temperature and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab28 (5 μg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 anti-mouse polyclonal antibody (ab96879) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.

  • developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 53 kDa

    All Lanes: Anti-p53 antibody [PAb 1801] (ab28) at 5 µg/ml

    Lane 1 and 10: Wild-type HCT116 cell lysate

    Lane 2 and 11: Wild-type HCT116 + irinotecan (10 µM, 24 hours) cell lysate

    Lane 3 and 12: p53 knockout HCT116 cell lysate

    Lane 4 and 13: p53 knockout HCT116 + irinotecan (10 µM, 24 hours) cell lysate

    Lane 5 and 14: A431 cell lysate (positive control)

    Lane 6 and 15: Saos-2 cell lysate (negative control)

    Lane 7 and 16: MEF cell lysate

    Lane 8 and 17: Wild-type HAP1 cell lysate

    Lane 9 and 18: p53 knockout HAP1 cell lysate

    Lanes 1-9: 1% BSA blocking buffer

    Lanes 10-18: 3% Milk blocking buffer

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

    We recommend using 3% milk as the blocking agent for western blot.

  • Overlay histogram showing HeLa cells stained with ab28 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween 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 (ab28, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

  • Analysis of recombinant p53 protein (ab43615) in MCF7 cells treated with vehicle (VEH)as well as MCF7 cells treated with camptothecin (CAM) and no material (NO) by sandwich ELISA with the use of anti-53 antibody (ab28) as capture and anti-p53 antibody (ab32389) as detector.
  • Anti-p53 antibody [PAb 1801] (ab28) at 5 µg/ml + MDA-MB-231 (Human breast adenocarcinoma cell line) Whole Cell Lysate at 20 µg

    Secondary
    Goat Anti-Mouse IgG H&L (HRP) preadsorbed (ab97040) at 1/10000 dilution
    developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 53 kDa
    Additional bands at : 53 kDa,60 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 20 minutes

    This blot was produced using a 10% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 3% Milk before being incubated with ab28 overnight at 4°C. Antibody binding was detected using an anti-mouse antibody conjugated to HRP, and visualised using ECL development solution.

     

    We recommend using 3% milk as the blocking agent for Western blot.



  • Predicted band size : 53 kDa

    The image shows a Western blot for ab28, testing the threshold detection of the p53 antibody. The lanes are as follows:

    Lane 1 - 20ug of A549 lysate, lane 2 - 10ug of A549 lysate, lane 3 - 5ug of A549 lysate, lane 4 - 2.5ug of A549 lysate, lane 5 - 1.5ug of A549 lysate.

    This picture was submitted as part of the review by Craig Carson.

     

References for Anti-p53 antibody [PAb 1801] (ab28)

This product has been referenced in:
  • Carrera S  et al. The role of the HIF-1a transcription factor in increased cell division at physiological oxygen tensions. PLoS One 9:e97938 (2014). WB ; Human . Read more (PubMed: 24835245) »
  • Stengel C  et al. In vivo and in vitro properties of STX2484: a novel non-steroidal anti-cancer compound active in taxane-resistant breast cancer. Br J Cancer 111:300-8 (2014). WB ; Human . Read more (PubMed: 24960406) »

See all 10 Publications for this product

Product Wall

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I am sorry about all the problems you have been having with ab28 in western blot. I just had one more protocol that you used.:

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Application Western blot
Sample Mouse Cell lysate - whole cell (Colon cancer cell line)
Gel Running Conditions Reduced Denaturing (12% gel)
Loading amount 30 µg
Treatment 10 µM CPT-11 for 24 hrs
Specification Colon cancer cell line
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Mrs. Anke Rauch

Verified customer

Submitted Jun 28 2016

Application Western blot
Sample Mouse Tissue lysate - whole (Colon lysate)
Gel Running Conditions Non-reduced Denaturing (10%)
Loading amount 20 µg
Treatment NoTreated and DSS-treated
Specification Colon lysate
Blocking step Milk as blocking agent for 20 minute(s) · Concentration: 10% · Temperature: 25°C
Username

Dr. Natalia Volodko

Verified customer

Submitted Aug 07 2015

Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (Human medulloblastoma cell line D556)
Permeabilization Yes - 0.3% Triton X-100
Specification Human medulloblastoma cell line D556
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted May 11 2015

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: 22°C
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: 10 mM Sodium Citrate
Sample Mouse Tissue sections (mouse brain cerebellum)
Specification mouse brain cerebellum
Permeabilization Yes - 0.3% Triton X-100
Fixative Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted Feb 23 2015

Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing
Sample Mouse Tissue lysate - whole (mouse cerebellum)
Specification mouse cerebellum
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted Apr 03 2014

Application Immunoprecipitation
Immuno-precipitation step Protein A/G
Sample Human Cell lysate - whole cell (Human 293T cells)
Specification Human 293T cells
Total protein in input 200 µg
Username

Abcam user community

Verified customer

Submitted Mar 31 2014

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