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  1. Link

    p53-antibody-do-1-chip-grade-ab1101.pdf

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Cell Biology Cell Cycle Cell Cycle Inhibitors p53
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Validated using a knockout cell line

Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)

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  • SDS
Reviews (13)Q&A (8)References (121)

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Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
  • Immunocytochemistry - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
  • Immunocytochemistry - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
  • ChIP - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
  • Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
  • Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
  • Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
  • Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
  • Flow Cytometry - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)

Key features and details

  • Mouse monoclonal [DO-1] to p53 - ChIP Grade
  • Suitable for: Flow Cyt, ICC, ChIP, IHC-P, WB
  • Knockout validated
  • Reacts with: Human
  • Isotype: IgG2a

Conjugates logo Related conjugates and formulations

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Overview

  • Product name

    Anti-p53 antibody [DO-1] - ChIP Grade
    See all p53 primary antibodies
  • Description

    Mouse monoclonal [DO-1] to p53 - ChIP Grade
  • Host species

    Mouse
  • Tested applications

    Suitable for: Flow Cyt, ICC, ChIP, IHC-P, WBmore details
  • Species reactivity

    Reacts with: Human
    Does not react with: Mouse, Rat
  • Immunogen

    Recombinant full length protein. This information is proprietary to Abcam and/or its suppliers.

  • Epitope

    The epitope maps to within aa 20-25.
  • Positive control

    • WB: A431, MCF7, HEK-293, DU145, MDA-MB-361 cell lysate. IHC-P: Human colon adenocarcinoma FFPE tissue sections. ICC: A431 cells. ICC KO: Hap1 cells (Hap1-p53 KO used as negative cell line) ChIP: HEK-293 cell chromatin. Flow Cyt: HeLa cells.
  • General notes

    This monoclonal p53 antibody has been knockout validated in Western blot. The expected band for p53 was observed in wild type HAP1 cell lysate and the band was not seen in TP53 knockout HAP1 cell lysate.

    This antibody clone is manufactured by Abcam. If you require a custom buffer formulation or conjugation for your experiments, please contact orders@abcam.com.

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.

    If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As

Properties

  • 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

    pH: 7.40
    Preservative: 0.02% Sodium azide
    Constituent: PBS

    Some batches contain 6.97% L-Arginine as a stabilizing agent. For lot-specific buffer information, please contact our Scientific Support team.
  • Concentration information loading...
  • Purity

    Protein G purified
  • Clonality

    Monoclonal
  • Clone number

    DO-1
  • Myeloma

    unknown
  • Isotype

    IgG2a
  • Light chain type

    kappa
  • Research areas

    • Cell Biology
    • Cell Cycle
    • Cell Cycle Inhibitors
    • p53
    • Cell Biology
    • Apoptosis
    • Intracellular
    • p53 Pathway
    • Epigenetics and Nuclear Signaling
    • DNA / RNA
    • DNA Damage & Repair
    • DNA Damage Response
    • p53
    • Epigenetics and Nuclear Signaling
    • Transcription
    • Cancer susceptibility
    • Tumor Suppressors
    • Epigenetics and Nuclear Signaling
    • Cell cycle
    • Cell Cycle Inhibitors
    • p53
    • Cancer
    • Cell cycle
    • Cell cycle inhibitors
    • p53 pathway
    • Cancer
    • Oncoproteins/suppressors
    • Tumor suppressors
    • p53 pathway
    • Neuroscience
    • Development
    • Neuroscience
    • Processes

Associated products

  • Alternative Versions

    • HRP Anti-p53 antibody [DO-1] (ab204452)
    • Anti-p53 antibody [DO-1] - BSA and Azide free (ab237976)
  • Assay kits

    • p53 Transcription Factor Assay Kit (Colorimetric) (ab207225)
  • ChIP Related Products

    • ChIP Kit (ab500)
  • Compatible Secondaries

    • Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) (ab150113)
    • Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150116)
    • Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772)
  • Isotype control

    • Mouse IgG2a, kappa monoclonal [MG2a-53] - Isotype control (ab18415)
  • Positive Controls

    • HeLa whole cell lysate (ab150035)
  • Recombinant Protein

    • Recombinant Human p53 protein (ab43615)
  • Related Products

    • Prestained Protein Ladder – Broad molecular weight (10-245 kDa) (ab116028)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab1101 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 (1)
Use at an assay dependent concentration.
ICC (1)
Use at an assay dependent concentration.
ChIP
Use at an assay dependent concentration.
IHC-P
Use a concentration of 1 - 5 µg/ml. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB (7)
Use at an assay dependent concentration. Predicted molecular weight: 43 kDa.
Notes
Flow Cyt
Use at an assay dependent concentration.
ICC
Use at an assay dependent concentration.
ChIP
Use at an assay dependent concentration.
IHC-P
Use a concentration of 1 - 5 µg/ml. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB
Use at an assay dependent concentration. Predicted molecular weight: 43 kDa.

Target

  • Function

    Acts 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 specificity

    Ubiquitous. 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 disease

    Note=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 similarities

    Belongs to the p53 family.
  • Domain

    The 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 localization

    Cytoplasm; 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.
  • Target information above from: UniProt accession P04637 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 7157 Human
    • Omim: 191170 Human
    • SwissProt: P04637 Human
    • Unigene: 654481 Human
    • 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 antigen p55 antibody
      • Tumor protein 53 antibody
      • Tumor protein p53 antibody
      • Tumor suppressor p53 antibody
      see all

    Images

    • Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      All lanes : Anti-p53 antibody [DO-1] - ChIP Grade (ab1101) at 1/1000 dilution

      Lane 1 : Wild-type A549 Vehicle Control Irinotecan (0 uM, 24 h) cell lysate
      Lane 2 : TP53 knockout A549 Vehicle Control Irinotecan (0 uM, 24 h) cell lysate
      Lane 3 : Wild-type Jurkat Vehicle Control Irinotecan (0 uM, 24 h) cell lysate
      Lane 4 : TP53 knockout Jurkat Vehicle Control Irinotecan (0 uM, 24 h) cell lysate
      Lane 5 : A431 cell lysate
      Lane 6 : Saos-2 cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 43 kDa
      Observed band size: 49 kDa why is the actual band size different from the predicted?



      False colour image of Western blot: Anti-p53 antibody [DO-1] - ChIP Grade staining at 1/1000 dilution, shown in green; Rabbit anti-alpha Tubulin antibody [EP1332Y] (ab52866) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab1101 was shown to bind specifically to p53. A band was observed at 49 kDa in wild-type A549 cell lysates with no signal observed at this size in tp53 knockout cell line ab276092 (knockout cell lysate ab282999). To generate this image, wild-type and tp53 knockout A549 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-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) at 1/20000 dilution.

    • Immunocytochemistry - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      Immunocytochemistry - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)

      ab1101 staining p53 - ChIP Grade in A431 cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab1101 at 5µg/ml and ab6046, Rabbit polyclonal to beta Tubulin - Loading Control. Cells were then incubated with ab150117, Goat polyclonal Secondary Antibody to Mouse IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150080, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 594) at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).

    • Immunocytochemistry - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      Immunocytochemistry - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)

      ab1101 staining p53 in wild-type Hap1 cells (top panel) and p53 knockout Hap1 cells (bottom panel). The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab1101 at 1µg/ml concentration and ab6046 (Rabbit polyclonal to beta Tubulin) at 1/1000 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to mouse IgG (Alexa Fluor® 488) (ab150117) at 2 μg/ml (shown in green) and a goat secondary antibody to rabbit IgG (Alexa Fluor® 594) (ab150080) at 2 μg/ml (shown in pseudo color red).  Nuclear DNA was labelled in blue with DAPI.

      Image was taken with a high-content analysis system (Perkin Elmer, Operetta CLS™).

    • ChIP - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      ChIP - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)

      Chromatin was prepared from HEK-293 (human epithelial cell line from embryonic kidney) cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25 µg of chromatin, 2 µg of ab1101, and 10 ml of protein A sepharose beads,10 ml of protein G sepharose beads. No IgG was added to the beads control. The immunoprecipitated DNA was quantified by real time PCR. Primers were as follows:

      Bax-1, forward: GGGTTATCTCTTGGGCTCACAA.

      Bax-1, reverse: GAGCTCTCCCCAGCGCA.

      Bax-2, forward: TGG AGC TGC AGA GGA TGA TTG

      Bax-2, reverse: CCA GTT GAA GTT GCC GTC AGA

      PUMA, forward: ATG CCT GCC TCA CCT TCA TC

      PUMA, reverse: TCA CAC GTC GCT CTC TCT AAA CC

      p21-1, forward: GCT GTG GCT CTG ATT GGC TTT

      p21-1, reverse: ACA GGC AGC CCA AGG ACA AA

      p21-2, forward: CAT CCC CAC AGC AGA GGA GAA

      p21-2, reverse: ACC CAG GCT TGG AGC AGC TA

      p21-3, forward: GAG TCC TGT TTG CTT CTG GGC A

      p21-3, reverse: CTG CAT TGG GGC TGC CTA TGT A

      PCNA, forward: CCA CCA TAA AGC TGG GGC TT

      PCNA, reverse: TCT CCC CGC CTC TTT GAC TC

    • Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      All lanes : Anti-p53 antibody [DO-1] - ChIP Grade (ab1101) at 0.4 µg/ml

      Lane 1 : Extract of HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) cells incubated with vehicle at 20 µg
      Lane 2 : Extract of HEK-293T cells incubated with etoposide at 20 µg
      Lane 3 : Extract of HEK-293T cells incubated with etoposide at 7.5 µg
      Lane 4 : Lambda phosphatase (diluted 1/400)-treated extract of HEK-293T cells incubated with etoposide at 7.5 µg
      Lane 5 : Lambda phosphatase (diluted 1/100)-treated extract of HEK-293T cells incubated with etoposide at 7.5 µg
      Lane 6 : Lambda phosphatase (diluted 1/25)-treated extract of HEK-293T cells incubated with etoposide at 7.5 µg
      Lane 7 : Extract of MCF7 (human breast adenocarcinoma cell line) cells incubated with vehicle at 20 µg
      Lane 8 : Extract of MCF7 cells incubated 6 hours with camptothecin at 20 µg
      Lane 9 : Extract of MCF7 cells incubated 16 hours with camptothecin at 20 µg
      Lane 10 : Extract of MCF7 cells incubated 24 hours with camptothecin at 20 µg

      Secondary
      All lanes : Goat anti mouse IgG(H&L)-HRP at 1/10000 dilution

      Predicted band size: 43 kDa
      Observed band size: 53 kDa why is the actual band size different from the predicted?



      Developed using the ECL technique with 30 second exposure.
      Blocking: 5% milk in PBS for 3 hours at RT.
      Primary antibody in 5% BSA + PBS was incubated overnight at 4°C.
      Secondary antibody in 5% milk + PBS was incubated for 2 hours at RT.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)

      IHC image of ab1101 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 (pH 6, epitope retrieval solution 1) for 20 minutes. The section was then incubated with ab1101, 1/500 dilution, for 15 minutes 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

    • Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      All lanes :

      Lanes 1 & 5 & 9 : Wild-type HAP1 cell lysate at 20 µg
      Lanes 2 & 6 & 10 : p53 knockout HAP1 cell lysate at 20 µg
      Lanes 3 & 7 : A431 cell lysate at 20 µg
      Lanes 4 & 8 & 12 : Saos-2 cell lysate at 20 µg
      Lane 11 : A431 cell lysate

      Predicted band size: 43 kDa



      Lanes 1, 2, 3 and 4: Green signal from target – ab1101 observed at 53 kDa
      Lanes 5, 6, 7 and 8: Red signal from loading control – ab181602 observed at 37 kDa
      Lanes 9, 10, 11 and 12: Merged (red and green) signal

      ab1101 was shown to specifically react with p53 in wild type HAP1 cells No band was observed in p53 knockout HAP1 cell lysates. Wild type and p53 knockout samples were subjected to SDS-PAGE. ab1101 and ab181602 (loading control to GAPDH) were diluted 1/1000 and 1/10 000 respectively and incubated overnight at 4ºC. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1/10 000 dilution for 1 h at room temperature before imaging.

    • Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      All lanes : Anti-p53 antibody [DO-1] - ChIP Grade (ab1101) at 1/1000 dilution

      Lane 1 : HEK-293 (human embryonic kidney cell line) whole cell lysate
      Lane 2 : DU 145 (human prostate carcinoma cell line) whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution

      Predicted band size: 43 kDa
      Observed band size: 50 kDa why is the actual band size different from the predicted?



      This blot was produced using a 4-12% 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 5% Milk before being incubated with ab1101 overnight at 4°C. Antibody binding was detected using ab175783 at a 1/10,000 dilution for 1 hour at room temperature and then imaged using the Licor Odyssey CLx.

    • Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      Western blot - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      All lanes : Anti-p53 antibody [DO-1] - ChIP Grade (ab1101) at 2.5 µg/ml

      Lane 1 : MDA-MB-361 (human breast adenocarcinoma cell line) whole cell lysate
      Lane 2 : MCF7 (human breast adenocarcinoma cell line) whole cell lysate
      Lane 3 : A549 (human lung adenocarcinoma epithelial cell line) whole cell lysate

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : Goat polyclonal to Mouse IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

      Predicted band size: 43 kDa
      Observed band size: 53 kDa why is the actual band size different from the predicted?
      Additional bands at: 55 kDa (possible post-translational modification)

    • Flow Cytometry - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)
      Flow Cytometry - Anti-p53 antibody [DO-1] - ChIP Grade (ab1101)

      Overlay histogram showing HeLa (human epithelial cell line from cervix adenocarcinoma) cells stained with ab1101 (red line). The cells were fixed with 4% paraformaldehyde for 10 minutes and then permeabilized with 0.1% PBS-Tween for 20 minutes. 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 (ab1101, 1/50 dilution) for 30 minutes at 22ºC. The secondary antibody used was a goat anti-mouse DyLight® 488 (IgG; H+L) ab96879 at 1/500 dilution for 30 minutes at 22ºC. Isotype control antibody (black line) was mouse IgG2a [ICIGG2A] ab91361, 1 µg/1 x 106 cells, used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HeLa cells fixed with 80% methanol (5 minutes)/permeabilized with 0.1% PBS-Tween for 20 minutes used under the same conditions.

    Protocols

    • ChIP protocols

    Click here to view the general protocols

    Datasheets and documents

    • SDS download

    • Datasheet download

      Download

    References (121)

    Publishing research using ab1101? Please let us know so that we can cite the reference in this datasheet.

    ab1101 has been referenced in 121 publications.

    • Kusumoto D  et al. Anti-senescent drug screening by deep learning-based morphology senescence scoring. Nat Commun 12:257 (2021). PubMed: 33431893
    • Xie L  et al. The HIF-1a/p53/miRNA-34a/Klotho axis in retinal pigment epithelial cells promotes subretinal fibrosis and exacerbates choroidal neovascularization. J Cell Mol Med 25:1700-1711 (2021). PubMed: 33438362
    • Charruyer A  et al. Decreased p53 is associated with a decline in asymmetric stem cell self-renewal in aged human epidermis. Aging Cell 20:e13310 (2021). PubMed: 33524216
    • Tang Q  et al. Mutant p53 regulates Survivin to foster lung metastasis. Genes Dev 35:528-541 (2021). PubMed: 33737385
    • Du J  et al. Pseudouridylate Synthase 7 Promotes Cell Proliferation and Invasion in Colon Cancer Through Activating PI3K/AKT/mTOR Signaling Pathway. Dig Dis Sci N/A:N/A (2021). PubMed: 33811565
    View all Publications for this product

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    Flow Cytometry abreview for Anti-p53 antibody [D01]

    Excellent
    Abreviews
    Abreviews
    Application
    Flow Cytometry
    Sample
    Human Cell (Human breast tumour)
    Specification
    Human breast tumour
    Preparation
    Cell harvesting/tissue preparation method: Trypsin/EDTA for 5 minutes
    Sample buffer: 1x PBS with 5% FCS
    Fixation
    70% ethanol
    Permeabilization
    No
    Gating Strategy
    live (non apoptotic cells) were selected
    Read More

    Dr. Penelope Ottewell

    Verified customer

    Submitted Nov 25 2008

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