Validated using a knockout cell line
Recombinant
RabMAb

Recombinant Anti-RUNX3 antibody [EPR20687] - BSA and Azide free (ab227125)

Overview

  • Product name

    Anti-RUNX3 antibody [EPR20687] - BSA and Azide free
    See all RUNX3 primary antibodies
  • Description

    Rabbit monoclonal [EPR20687] to RUNX3 - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

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

    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human RUNX3 aa 150-250. The exact sequence is proprietary.
    Database link: Q13761

  • Positive control

    • IHC: Human stomach tissue.
  • General notes

    Ab227125 is the carrier-free version of ab224641. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    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.

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

    Constituent: 59% PBS
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR20687
  • Isotype

    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab227125 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.
IP Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration. 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. Detects a band of approximately 44, 46 kDa (predicted molecular weight: 44 kDa).
ICC/IF Use at an assay dependent concentration.

Target

  • Function

    CBF binds to the core site, 5'-PYGPYGGT-3', of a number of enhancers and promoters, including murine leukemia virus, polyomavirus enhancer, T-cell receptor enhancers, lck, IL-3 and GM-CSF promoters.
  • Sequence similarities

    Contains 1 Runt domain.
  • Domain

    A proline/serine/threonine rich region at the C-terminus is necessary for transcriptional activation of target genes.
  • Post-translational
    modifications

    Phosphorylated on tyrosine residues by SRC. Phosphorylated by LCK and FYN.
  • Cellular localization

    Nucleus. Cytoplasm. The tyrosine phosphorylated form localizes to the cytoplasm.
  • Information by UniProt
  • Database links

  • Alternative names

    • Acute myeloid leukemia 2 protein antibody
    • Acute myeloid leukemia gene 2 antibody
    • AML 2 antibody
    • AML2 antibody
    • CBF alpha 3 antibody
    • CBF-alpha-3 antibody
    • CBFA 3 antibody
    • CBFA3 antibody
    • Core binding factor alpha 3 subunit antibody
    • core binding factor antibody
    • Core binding factor runt domain alpha subunit 3 antibody
    • Core binding factor subunit alpha 3 antibody
    • core-binding factor antibody
    • Core-binding factor subunit alpha-3 antibody
    • Oncogene AML 2 antibody
    • Oncogene AML-2 antibody
    • PEA2 alpha C antibody
    • PEA2-alpha C antibody
    • PEBP2 alpha C antibody
    • PEBP2-alpha C antibody
    • Pebp2a3 antibody
    • PEBP2aC antibody
    • Polyomavirus enhancer binding protein 2 alpha C subunit antibody
    • Polyomavirus enhancer-binding protein 2 alpha C subunit antibody
    • runt domain alpha subunit 3 antibody
    • runt related transcription factor 3 antibody
    • Runt-related transcription factor 3 antibody
    • RUNX 3 antibody
    • Runx3 antibody
    • RUNX3_HUMAN antibody
    • SL3 3 enhancer factor 1 alpha C subunit antibody
    • SL3-3 enhancer factor 1 alpha C subunit antibody
    • SL3/AKV core binding factor alpha C subunit antibody
    • SL3/AKV core-binding factor alpha C subunit antibody
    • Transcription factor AML2 antibody
    see all

Images

  • All lanes : Anti-RUNX3 antibody [EPR20687] (ab224641) at 1/1000 dilution

    Lane 1 : Wild-type HAP1 whole cell lysate
    Lane 2 : RUNX3 knockout HAP1 whole cell lysate
    Lane 3 : Ramos whole cell lysate
    Lane 4 : HeLa whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 44 kDa



    Lanes 1 - 4: Merged signal (red and green). Green - ab224641 observed at 44-46 kDa. Red - loading control, ab130007, observed at 130 kDa.

    ab224641 was shown to specifically react with RUNX3 in wild-type HAP1 cells as signal was lost in RUNX3 knockout cells. Wild-type and RUNX3 knockout samples were subjected to SDS-PAGE. Ab224641 and ab130007 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab224641).

  • RUNX3 was immunoprecipitated from 0.35 mg of Raji (human Burkitt's lymphoma cell line) whole cell lysate with ab224641 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab224641 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution.

    Lane 1: Raji whole cell lysate 10 µg (Input). 

    Lane 2: ab224641 IP in Raji whole cell lysate (+). 

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab224641 in Raji whole cell lysate (-).

    Blocking and dilution 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 (ab224641).

  • Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol permeabilized Raji (human Burkitt's lymphoma cell line) cell line labeling RUNX3 with ab224641 at 1/500 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab224641).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized Raji (human Burkitt's lymphoma cell line) cells labeling RUNX3 with ab224641 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on Raji cell line. DAPI (blue) and anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution were used as counterstains.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is AlexaFluor®488 Goat anti-Rabbit (ab150077) at 1/1000 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab224641).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized Ramos (human Burkitt's lymphoma cell line) cells labeling RUNX3 with ab224641 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear staining on Ramos cell line. DAPI (blue) and anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution were used as counterstains.

    The negative controls are as follows:

    Negative control: MCF7 (human breast adenocarcinoma cell line) (PMID: 21706051).

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is AlexaFluor®488 Goat anti-Rabbit (ab150077) at 1/1000 dilution..

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab224641).

  • Chromatin was prepared from U-937 (PMA treated or not) cells according to the Abcam X-ChIP protocol. Cells were fixed with 1% formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 5µg of ab224641 (red), and 20µl of protein A/G sepharose beads slurry (10µl of sepharose A beads + 10µl of sepharose G beads). 5μg of rabbit normal IgG was added to the beads control (grey). The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab224641).

     

  • Immunohistochemical analysis of paraffin-embedded human diffuse large B cell lymphoma tissue labeling RUNX3 with ab224641 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051), ready to use. Nuclear staining on human diffuse large B cell lymphoma (PMID:27184221) is observed. 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), ready to use.

    Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab224641).

  • Immunohistochemical analysis of paraffin-embedded human gastric cancer tissue labeling RUNX3 with ab224641 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051), ready to use. Nuclear staining on lymphoid cells of human gastric cancer (PMID:27566570) is observed. 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), ready to use.

    Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab224641).

  • Immunohistochemical analysis of paraffin-embedded human stomach tissue labeling RUNX3 with ab224641 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051), ready to use. Nuclear staining on lymphoid cells of human stomach (PMID:15514019; PMID:21786422) is observed. 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), ready to use.

    Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab224641).

References

ab227125 has not yet been referenced specifically in any publications.

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