Overview

  • Product name

    Anti-KDM5A / Jarid1A / RBBP2 antibody [EPR18651]
    See all KDM5A / Jarid1A / RBBP2 primary antibodies
  • Description

    Rabbit monoclonal [EPR18651] to KDM5A / Jarid1A / RBBP2
  • Host species

    Rabbit
  • Tested applications

    Suitable for: ICC/IF, IP, WB, Flow Cytmore details
  • Species reactivity

    Reacts with: Mouse, Human
  • Immunogen

    Recombinant fragment within Human KDM5A/ Jarid1A/ RBBP2 aa 1450 to the C-terminus. The exact sequence is proprietary.
    Database link: P29375

  • Positive control

    • WB: HeLa, LCC, HEK-293 and NIH/3T3 whole cell lysates; mouse testis lysate. ICC/IF: HeLa cells. IP: HeLa whole cell lysate. FC: NIH/3T3 cell lysate
  • 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.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab194286 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
ICC/IF 1/2000.
IP 1/100.
WB 1/5000. Detects a band of approximately 192 kDa (predicted molecular weight: 192 kDa).
Flow Cyt 1/1500.

Target

  • Function

    Histone demethylase that specifically demethylates 'Lys-4' of histone H3, thereby playing a central role in histone code. Does not demethylate histone H3 'Lys-9', H3 'Lys-27', H3 'Lys-36', H3 'Lys-79' or H4 'Lys-20'. Demethylates trimethylated and dimethylated but not monomethylated H3 'Lys-4'. May stimulate transcription mediated by nuclear receptors. May be involved in transcriptional regulation of Hox proteins during cell differentiation. May participate in transcriptional repression of cytokines such as CXCL12.
  • Sequence similarities

    Belongs to the JARID1 histone demethylase family.
    Contains 1 ARID domain.
    Contains 1 JmjC domain.
    Contains 1 JmjN domain.
    Contains 3 PHD-type zinc fingers.
  • Domain

    The GSGFP motif is required for the interaction with SUZ12.
  • Cellular localization

    Nucleus > nucleolus. Occupies promoters of genes involved in RNA metabolism and mitochondrial function.
  • Information by UniProt
  • Database links

  • Alternative names

    • Histone demethylase JARID1A antibody
    • JARID1A antibody
    • Jumonji/ARID domain containing protein 1A antibody
    • Jumonji/ARID domain-containing protein 1A antibody
    • Kdm5a antibody
    • KDM5A_HUMAN antibody
    • Lysine-specific demethylase 5A antibody
    • RBBP-2 antibody
    • RBBP2 antibody
    • RBP2 antibody
    • Retinoblastoma binding protein 2 antibody
    • Retinoblastoma-binding protein 2 antibody
    see all

Images

  • Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
    Lane 2: KDM5A knockout HAP1 whole cell lysate (20 µg)
    Lane 3: HeLa whole cell lysate (20 µg)
    Lane 4: HEK293 whole cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab194286 observed at 240 kDa. Red - loading control, ab18058, observed at 130 kDa.

    ab194286 was shown to specifically recognize KDM5A in wild-type HAP1 cells along with additional cross-reactive bands. No band was observed when KDM5A knockout samples were examined. Wild-type and KDM5A knockout samples were subjected to SDS-PAGE. Ab194286 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 1/2500 dilution and 1/10000 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.

     

     

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling KDM5A / Jarid1A / RBBP2 with ab194286 at 1/2000 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 HeLa cell line. The nuclear counter stain is DAPI (blue).

    Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution (red).

    The negative controls are as follows:
    -ve control 1: ab194286 at 1/2000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary at 1/1000 dilution.
    -ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary at 1/1000 dilution.

  • All lanes : Anti-KDM5A / Jarid1A / RBBP2 antibody [EPR18651] (ab194286) at 1/5000 dilution

    Lane 1 : LLC (Mouse lung carcinoma) whole cell lysate
    Lane 2 : Mouse testis lysate
    Lane 3 : HEK-293 (Human epithelial cells from embryonic kidney) whole cell lysate

    Lysates/proteins at 20 µg per lane.

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

    Predicted band size: 192 kDa
    Observed band size: 192 kDa


    Exposure time: 30 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Anti-KDM5A / Jarid1A / RBBP2 antibody [EPR18651] (ab194286) at 1/5000 dilution + HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate at 20 µg

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

    Predicted band size: 192 kDa
    Observed band size: 192 kDa


    Exposure time: 10 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Anti-KDM5A / Jarid1A / RBBP2 antibody [EPR18651] (ab194286) at 1/1000 dilution + NIH/3T3 (Mouse embryonic fibroblast cells) whole cell lysate at 10 µg

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

    Predicted band size: 192 kDa
    Observed band size: 192 kDa


    Exposure time: 10 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • KDM5A / Jarid1A / RBBP2 was immunoprecipitated from 1mg of HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysate with ab194286 at 1/100 dilution. Western blot was performed from the immunoprecipitate using ab194286 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

    Lane 1: HeLa whole cell lysate 10ug (Input).

    Lane 2: ab194286 IP in HeLa whole cell lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab194286 in HeLa whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 5 seconds.

  • Flow cytometry analysis of NIH/3T3 (mouse embryo) cells labelling KDM5A / Jarid1A / RBBP2 (red) with purified ab194286 at dilution of 1/1500. The secondary antibody used was Alexa Fluor® 488 goat-anti-rabbit IgG (1/2000). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Isotype control antibody used was Rabbit Monoclonal IgG (black). The blue line shows cells without incubation with primary antibody and secondary antibody.

References

This product has been referenced in:

  • McBrayer SK  et al. Autochthonous tumors driven by Rb1 loss have an ongoing requirement for the RBP2 histone demethylase. Proc Natl Acad Sci U S A N/A:N/A (2018). Read more (PubMed: 29610306) »
See 1 Publication for this product

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