Overview

  • Product name
    Anti-CTCF antibody [EPR7314(B)]
    See all CTCF primary antibodies
  • Description
    Rabbit monoclonal [EPR7314(B)] to CTCF
  • Host species
    Rabbit
  • Tested applications
    Suitable for: ChIP, WB, IHC-P, Flow Cyt, ICC/IFmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human CTCF aa 700-800. The exact sequence is proprietary.
    (Peptide available as ab209492)

  • Positive control
    • WB- HeLa and 293T cell lysates, Human colon tissue, Mouse brain tissue and rat heart tissue. IHC- Human breast carcinoma, mouse and rat kideny ICC/IF- HeLa cell lysates Flow cytometry- 293T cell lysates
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab128873 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.
WB 1/1000 - 1/10000. Detects a band of approximately 140 kDa (predicted molecular weight: 83 kDa).Can be blocked with CTCF peptide (ab209492).
IHC-P 1/250 - 1/1000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Flow Cyt 1/40 - 1/1000.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

 

ICC/IF 1/250 - 1/500.

Target

  • Function
    Chromatin binding factor that binds to DNA sequence specific sites. Involved in transcriptional regulation by binding to chromatin insulators and preventing interaction between promoter and nearby enhancers and silencers. Acts as transcriptional repressor binding to promoters of vertebrate MYC gene and BAG1 gene. Also binds to the PLK and PIM1 promoters. Acts as a transcriptional activator of APP. Regulates APOA1/C3/A4/A5 gene cluster and controls MHC class II gene expression. Plays an essential role in oocyte and preimplantation embryo development by activating or repressing transcription. Seems to act as tumor suppressor. Plays a critical role in the epigenetic regulation. Participates to the allele-specific gene expression at the imprinted IGF2/H19 gene locus. On the maternal allele, binding within the H19 imprinting control region (ICR) mediates maternally inherited higher-order chromatin conformation to restrict enhancer access to IGF2. Plays a critical role in gene silencing over considerable distances in the genome. Preferentially interacts with unmethylated DNA, preventing spreading of CpG methylation and maintaining methylation-free zones. Inversely, binding to target sites is prevented by CpG methylation. Plays a important role in chromatin remodeling. Can dimerize when it is bound to different DNA sequences, mediating long-range chromatin looping. Mediates interchromosomal association between IGF2/H19 and WSB1/NF1 and may direct distant DNA segments to a common transcription factory. Causes local loss of histone acetylation and gain of histone methylation in the beta-globin locus, without affecting transcription. When bound to chromatin, it provides an anchor point for nucleosomes positioning. Seems to be essential for homologous X-chromosome pairing. May participate with Tsix in establishing a regulatable epigenetic switch for X chromosome inactivation. May play a role in preventing the propagation of stable methylation at the escape genes from X- inactivation. Involved in sister chromatid cohesion. Associates with both centromeres and chromosomal arms during metaphase and required for cohesin localization to CTCF sites. Regulates asynchronous replication of IGF2/H19.
  • Tissue specificity
    Ubiquitous. Absent in primary spermatocytes.
  • Sequence similarities
    Belongs to the CTCF zinc-finger protein family.
    Contains 11 C2H2-type zinc fingers.
  • Domain
    The 11 zinc fingers are highly conserved among vertebrates, exhibiting almost identical amino acid sequences. Different subsets or combination of individual zinc fingers gives the ability to CTCF to recognize multiple DNA target sites.
  • Post-translational
    modifications
    Sumoylated on Lys-74 and Lys-689; sumoylation of CTCF contributes to the repressive function of CTCF on the MYC P2 promoter.
  • Cellular localization
    Nucleus > nucleoplasm. Chromosome. Chromosome > centromere. May translocate to the nucleolus upon cell differentiation. Associates with both centromeres and chromosomal arms during metaphase. Associates with the H19 ICR in mitotic chromosomes. May be preferentially excluded from heterochromatin during interphase.
  • Information by UniProt
  • Database links
  • Alternative names
    • 11 zinc finger protein antibody
    • 11 zinc finger transcriptional repressor antibody
    • 11-zinc finger protein antibody
    • CCCTC binding factor (zinc finger protein) antibody
    • CCCTC binding factor antibody
    • CCCTC-binding factor antibody
    • Ctcf antibody
    • CTCF_HUMAN antibody
    • CTCFL paralog antibody
    • MRD21 antibody
    • Transcriptional repressor CTCF antibody
    see all

Images

  • All lanes : Anti-CTCF antibody [EPR7314(B)] (ab128873) at 1/10000 dilution

    Lane 1 : HeLa (human cervix adenocarcinoma) whole cell lysate
    Lane 2 : 293T (human embryonic kidney) whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 83 kDa
    Observed band size: 140 kDa
    why is the actual band size different from the predicted?



    Blocking and diluting buffer 5% NFDM/TBST
  • Anti-CTCF antibody [EPR7314(B)] (ab128873) at 1/10000 dilution + Mouse brain at 20 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 83 kDa
    Observed band size: 140 kDa why is the actual band size different from the predicted?



    Blocking and diluting buffer 5% NFDM/TBST
  • Anti-CTCF antibody [EPR7314(B)] (ab128873) at 1/10000 dilution + Rat heart at 20 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 83 kDa
    Observed band size: 140 kDa why is the actual band size different from the predicted?



    Blocking and diluting buffer 5% NFDM/TBST
  • Chromatin was prepared from HeLa 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 ab128873 (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).

  • All lanes : Anti-CTCF antibody [EPR7314(B)] (ab128873) at 1/1000 dilution (un-purified)

    Lane 1 : HeLa cell lysate
    Lane 2 : 293T cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat anti-Rabbit HRP at 1/2000 dilution

    Predicted band size: 83 kDa

  • Immunohistochemical staining of paraffin-embedded human breast carcinoma sections labelling CTCF with purified ab128873 at dilution of 1:1000. The secondary antibody used was ab97051; a goat anti-rabbit IgG H&L (HRP) at dilution of 1/500. The sample was counter-stained with hematoxylin. Antigen retrieval was performed using EDTA Buffer; pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

  • Immunohistochemical staining of paraffin-embedded mouse kidney sections labelling CTCF with purified ab128873 at dilution of 1:1000. The secondary antibody used was ab97051; a goat anti-rabbit IgG H&L (HRP) at dilution of 1/500. The sample was counter-stained with hematoxylin. Antigen retrieval was performed using EDTA Buffer; pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

  • Immunohistochemical staining of paraffin-embedded rat kidney sections labelling CTCF with purified ab128873 at dilution of 1:1000. The secondary antibody used was ab97051; a goat anti-rabbit IgG H&L (HRP) at dilution of 1/500. The sample was counter-stained with hematoxylin. Antigen retrieval was performed using EDTA Buffer; pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

  • Immunohistochemical analysis of paraffin embedded Human colon tissue labelling CTCF with un-purified ab128873 at 1/250 dilution.

  • Immunocytochemistry/immunofluorescence staining of 4% paraformaldehyde fixed; 0.1% triton X 100 permeabilized HeLa (human cervix adenocarcinoma) cells with purified ab128873 at dilution of 1/500. The secondary antibody used was Alexa Fluor® 488; goat anti-rabbit IgG (ab150077) at a dilution of 1/1000. Nucleus was counter-stained with DAPI (blue). ab7291, a mouse anti-tubulin antibody (1/1000) was used to stain tubulin along with ab150120 (AlexaFluor®594 goat anti-mouse secondary, 1/1000) shown in the top right hand panel. The negative controls are shown in the bottom middle and right hand panels- for negative control 1 rabbit primary antibody and anti-mouse secondary antibody (ab150120) was used. For negative control 2 mouse primary antibody (ab7291) and anti-rabbit secondary antibody (ab150077) was used.

  • Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling CTCF with un-purified ab128873 at 1/250 dilution.

  • Flow cytometry analysis showing 4% paraformaldehyde fixed 293T (human embryonic kidney) cells labelling CTCF with purified ab128873 at dilution of 1/40 followed by the secondary antibody; Alexa Fluor® 488 goat-anti-rabbit IgG at dilution of 1/500 (red line). A non-specific IgG antibody (rabbit monoclonal) was used as isotype control (black line). The blue line shows cells without incubation with primary antibody and secondary antibody.

  • Overlay histogram showing Hela cells stained with un-purified ab128873 (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 (ab128873, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/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 antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

  • Equilibrium disassociation constant (KD)
    Learn more about KD

    Click here to learn more about KD

References

This product has been referenced in:
  • Ginno PA  et al. Cell cycle-resolved chromatin proteomics reveals the extent of mitotic preservation of the genomic regulatory landscape. Nat Commun 9:4048 (2018). Read more (PubMed: 30279501) »
  • Hansen AS  et al. CTCF and cohesin regulate chromatin loop stability with distinct dynamics. Elife 6:N/A (2017). Read more (PubMed: 28467304) »
See all 4 Publications for this product

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