Overview

  • Product nameAnti-PCNA [PC10] antibodySee all PCNA primary antibodies ...
  • Description
    Mouse monoclonal [PC10] to PCNA
  • SpecificityThis antibody is specific for PCNA p36 protein expressed at high levels in proliferating cells.
  • Tested applicationsIHC-P, IP, WB, ICC/IF, IHC-Fr, Flow Cyt more details
  • Species reactivity
    Reacts with: Mouse, Rat, Chicken, Human, Pig, Fruit fly (Drosophila melanogaster), Monkey, Zebrafish, Dogfish/Catshark, Thornback ray
    Predicted to work with: Cow
  • Immunogen

    Protein A-rat PCNA (proliferating cell nuclear antigen) fusion obtained from pC2T

  • Positive control
    • WB: DT40 B lymphoma cell lysate, 293 cell lysate (see review), Hela, HEK293, A431 whole cell lysates. IHC-P: mouse hippocampus (see review), Normal human tonsil, developing chick brain. IHC-Fr: rat intestine. Flow Cyt: HeLa.

Properties

Applications

Our Abpromise guarantee covers the use of ab29 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Notes
IHC-P 1/10000 - 1/30000. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
IP Use at an assay dependent concentration.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 30 kDa (predicted molecular weight: 29 kDa).
ICC/IF Use a concentration of 0.5 - 1 µg/ml. Methanol fixation recommended
IHC-Fr Use at an assay dependent concentration.
Flow Cyt Use 1µg for 106 cells.

Target

  • FunctionThis protein is an auxiliary protein of DNA polymerase delta and is involved in the control of eukaryotic DNA replication by increasing the polymerase's processibility during elongation of the leading strand. Induces a robust stimulatory effect on the 3'-5' exonuclease and 3'-phosphodiesterase, but not apurinic-apyrimidinic (AP) endonuclease, APEX2 activities. Has to be loaded onto DNA in order to be able to stimulate APEX2.
  • Sequence similaritiesBelongs to the PCNA family.
  • Post-translational
    modifications
    Upon methyl methanesulfonate-induced DNA damage, mono-ubiquitinated by the UBE2B-RAD18 complex on Lys-164. This induces non-canonical polyubiquitination on Lys-164 through 'Lys-63' linkage of ubiquitin moieties by the E2 complex UBE2N-UBE2V2 and the E3 ligases, HLTF, RNF8 and SHPRH, which is required for DNA repair. 'Lys-63' polyubiquitination prevents genomic instability on DNA damage. Monoubiquitination at Lys-164 also takes place in undamaged proliferating cells, and is mediated by the DCX(DTL) complex, leading to enhance PCNA-dependent translesion DNA synthesis.
    Acetylated in response to UV irradiation. Acetylation disrupts interaction with NUDT15 and promotes degradation.
  • Cellular localizationNucleus. Forms nuclear foci representing sites of ongoing DNA replication and vary in morphology and number during S phase. Together with APEX2, is redistributed in discrete nuclear foci in presence of oxidative DNA damaging agents.
  • Target information above from: UniProt accession P12004 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links
    see all
  • Alternative names
    • Cyclin antibody
    • DNA polymerase delta auxiliary protein antibody
    • HGCN8729 antibody
    • MGC8367 antibody
    • Mutagen-sensitive 209 protein antibody
    • OTTHUMP00000030189 antibody
    • OTTHUMP00000030190 antibody
    • PCNA antibody
    • Pcna/cyclin antibody
    • PCNA_HUMAN antibody
    • PCNAR antibody
    • Polymerase delta accessory protein antibody
    • Proliferating Cell Nuclear Antigen antibody
    see all

Anti-PCNA [PC10] antibody images

  • IHC image of PCNA staining in rat spleen 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 ab29, 1/30,000 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.

    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.

  • IHC image of PCNA staining in rat large intestine 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 ab29, 0.025µg/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.

  • Mouse monoclonal [PC10] to PCNA - Proliferation Marker (ab29) used in immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections;1/6000 for 2h at RT) on intestine of adult Zebra fish). Antigen retrieval step: Heat mediated. Blocking step: 1% BSA for 10 mins at RT. Incubation time: Secondary Antibody: Biotin conjugated goat anti mouse Igs (1/200). NB: The crypt nuclei on this image of zebrafish intestine, are positive for the PCNA/PC10 clone conforming to accepted localisation data for PCNA in other species.
  • Mouse monoclonal [PC10] to PCNA - Proliferation Marker (ab29) used in immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections; 1/6000 for 2h at RT) on Human Tissue sections (Paget's disease of the nipple). Antigen retrieval step: Heat mediated. Blocking step: 1% BSA for 10 mins at RT. Incubation time: Secondary Antibody: Biotin conjugated goat anti mouse Igs (1/200).
  • All lanes : Anti-PCNA [PC10] antibody (ab29) at 5 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : HEK293 (Human embryonic kidney cell line) Whole Cell Lysate
    Lane 3 : A431 (Human epithelial carcinoma cell line) Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

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

    Performed under reducing conditions.

    Predicted band size : 29 kDa
    Observed band size : 29 kDa


    Exposure time : 4 minutes
  • Overlay histogram showing HeLa cells stained with ab29 (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 (ab29, 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 IgG2a [ICIGG2A] (ab91361, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

  • All lanes : Anti-PCNA [PC10] antibody (ab29) at 1/5000 dilution

    Lane 1 : Human 293 total cell extract
    Lane 2 : Chicken DT40 total cell extract


    Performed under reducing conditions.

    Predicted band size : 29 kDa
    Observed band size : 30 kDa (why is the actual band size different from the predicted?)


    Exposure time : 5 seconds

    Taken from an Abreview submitted by Javier di Noia

    Blocking for 2 hours in 5% milk. Primary antibody incubated for 1 hour.
  • ab29 at a 1/500 dilution used in immunoprecipitation.

    Lane 1: Beads only + HeLa whole cell lysate

    Lane 2: Beads only + SupT1 whole cell lysate

    Lane 3: Beads + ab29 + HeLa whole cell lysate

    Lane 4: Beads + ab29 + SupT1 whole cell lysate

    A 50KDa band is precipitated which is IgG heavy chain whilst the 30kDa band is PCNA.

    See Abreview

  • ab29 at 1/6000 staining mouse embryo (day 17) liver and gut tissue sections by IHC-P. The tissue was formaldehyde fixed and a heat mediated antigen retrieval step in Tris buffer was performed. The tissue was blocked before incubation with the antibody for 2 hours. A biotinylated goat polyclonal antibody was used as the secondary.

    See Abreview

  • Mouse monoclonal [PC10] to PCNA - Proliferation Marker (ab29) used in immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections; 1/6000 for 2h at RT) on E6 developing chick brain). Antigen retrieval step: Heat mediated. Blocking step: 1% BSA for 10 mins at RT. Incubation time: Secondary Antibody: Biotin conjugated goat anti mouse Igs (1/200). NB: This image shows developing brain/overlying skin.
  • Immunohistochemistical staining (Formaldehyde/PFA-fixed paraffin-embedded sections) for PCNA antibody [PC10] - Proliferation Marker (ab29) on Rat Tissue sections (adult spinal cord DRG). Antigen retrieval step: Heat mediated. Blocking step: 1% BSA for 10 mins at RT. Primary Antibody used at 1/6000 for 2 minutes at RT. Secondary Antibody: Biotin labelled goat anti mouse Igs (1/200).
  • ab29 staining PCNA in mouse embryonic brain tissue section by Immunohistochemistry (Frozen sections). Tissue samples were fixed with paraformaldehyde and permeabilized with 0.1% PBS-Tween before blocking with 5% BSA for 1 hour at 220C. The sample was incubated with primary antibody (1/500) in 5% BSA in 0.3% PBS-Triton-X100 for 14 hours at 220C. An Alexa Fluor®488-conjugated Goat polyclonal to mouse IgG was used as secondary antibody at 1/500 dilution.

    See Abreview

  • ICC/IF image of ab29 stained HeLa cells. The cells were 100% methanol fixed (5 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 (ab29, 5µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-mouse IgG - H&L, pre-adsorbed (ab96879) used at a 1/250 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 for Anti-PCNA [PC10] antibody (ab29)

This product has been referenced in:
  • Lim S  et al. SNAI1-mediated epithelial-mesenchymal transition confers chemoresistance and cellular plasticity by regulating genes involved in cell death and stem cell maintenance. PLoS One 8:e66558 (2013). WB ; Human . Read more (PubMed: 23799116) »
  • Vera A  et al. SCO-spondin from embryonic cerebrospinal fluid is required for neurogenesis during early brain development. Front Cell Neurosci 7:80 (2013). Chicken . Read more (PubMed: 23761733) »

See all 51 Publications for this product

Product Wall



The immunogen used to produce ab29 is full length rat PCNA fused to Protein A. The exact epitope recognized by ab29 has not been determined.

Application Flow Cytometry
Fixation Paraformaldehyde
Permeabilization Yes - 0.1% Triton in PBS
Sample Human Cell (MCF-7 (invasive ductal carcinoma line))
Specification MCF-7 (invasive ductal carcinoma line)
Gating Strategy isotype control (black line in histogram)
Preparation Cell harvesting/tissue preparation method: accutase and accumax dissociation
Sample buffer: PBS 7.4 + 10% FBS for washing
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Submitted Mar 12 2014

Application Immunocytochemistry/ Immunofluorescence
Blocking step Dako Protein Block as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 20°C
Sample Pig Cell (Intestinal epithelium)
Specification Intestinal epithelium
Permeabilization Yes - 0.3% PBS-T
Fixative Paraformaldehyde
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Dr. Liara Gonzalez

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Submitted Feb 05 2014

Application Western blot
Loading amount 25 µg
Gel Running Conditions Reduced Denaturing (4-12%)
Sample Human Cell lysate - whole cell (HCT116)
Specification HCT116
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
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Submitted Dec 19 2013

Application Immunocytochemistry/ Immunofluorescence
Blocking step Serum as blocking agent for 10 minute(s) · Concentration: 2% · Temperature: 25°C
Sample Rat Cell (Oliogdendorcytes)
Specification Oliogdendorcytes
Permeabilization Yes - 0.01% Triton X100
Fixative Paraformaldehyde
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Submitted Dec 10 2013

Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step 2% normal goat serum, 1% bovine serum albumin, 0.1% cold fish skin gelatine, 0.1% Triton X-100, 0.05% Tween 20, 0.05% sodium azide, 0.01M PBS, pH 7.2 as blocking agent for 30 minute(s) · Concentration: 2% · Temperature: 25°C
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: Citrate Buffer (pH 9.0)
Sample Frog Tissue sections (Proximal Small intestine)
Specification Proximal Small intestine
Permeabilization No
Fixative Formaldehyde
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Dr. Rebecca Cramp

Verified customer

Submitted Nov 22 2013

Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Antigen retrieval step Heat mediated
Sample hairy pufferfish Tissue sections (epithelial cells)
Specification epithelial cells
Permeabilization No
Fixative Paraformaldehyde
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Mr. Liam Rasch

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Submitted Nov 04 2013

Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Antigen retrieval step Heat mediated
Sample Thornback ray Tissue sections (eye cells)
Specification eye cells
Fixative Paraformaldehyde
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Mr. Liam Rasch

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Submitted Oct 01 2013

Application Western blot
Sample Human Cell lysate - whole cell (human colon cancer cells)
Loading amount 40 µg
Specification human colon cancer cells
Gel Running Conditions Reduced Denaturing (4-12%)
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
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Submitted Apr 22 2013

Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample Fish Tissue sections (oral epithelium)
Specification oral epithelium
Fixative Paraformaldehyde
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: Citric acid buffer pH6
Permeabilization No
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Submitted Apr 18 2013

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