Validated using a knockout cell line
Recombinant
RabMAb

Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade (ab191250)

Overview

  • Product name
    Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade
    See all KMT6 / EZH2 primary antibodies
  • Description
    Rabbit monoclonal [EPR20108] to KMT6 / EZH2 - ChIP Grade
  • Tested applications
    Suitable for: WB, ChIP, IPmore details
  • Species reactivity
    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human KMT6/ EZH2 aa 350-450. The exact sequence is proprietary.
    Database link: Q15910

  • Positive control
    • WB: MCF7, HeLa, U-2 OS and HEK-293 whole cell lysates; Wild-type HAP1 cell lysate. IP: HeLa whole cell lysate. ChIP: Chromatin from MDA-MB-231 cells.
  • 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 ab191250 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
WB 1/1000. Detects a band of approximately 90 kDa (predicted molecular weight: 85 kDa).
ChIP Use 5 µg for 25 µg of chromatin.
IP 1/30.

Target

  • Function
    Polycomb group (PcG) protein. Catalytic subunit of the PRC2/EED-EZH2 complex, which methylates 'Lys-9' and 'Lys-27' of histone H3, leading to transcriptional repression of the affected target gene. Able to mono-, di- and trimethylate 'Lys-27' of histone H3 to form H3K27me1, H3K27me2 and H3K27me3, respectively. Compared to EZH2-containing complexes, it is more abundant in embryonic stem cells and plays a major role in forming H3K27me3, which is required for embryonic stem cell identity and proper differentiation. The PRC2/EED-EZH2 complex may also serve as a recruiting platform for DNA methyltransferases, thereby linking two epigenetic repression systems. Genes repressed by the PRC2/EED-EZH2 complex include HOXC8, HOXA9, MYT1, CDKN2A and retinoic acid target genes.
  • Tissue specificity
    Expressed in many tissues. Overexpressed in numerous tumor types including carcinomas of the breast, colon, larynx, lymphoma and testis.
  • Sequence similarities
    Belongs to the histone-lysine methyltransferase family. EZ subfamily.
    Contains 1 SET domain.
  • Developmental stage
    Expression decreases during senescence of embryonic fibroblasts (HEFs). Expression peaks at the G1/S phase boundary.
  • Post-translational
    modifications
    Phosphorylated by AKT1. Phosphorylation by AKT1 reduces methyltransferase activity.
  • Cellular localization
    Nucleus.
  • Information by UniProt
  • Database links
  • Alternative names
    • Enhancer of zeste 2 antibody
    • enhancer of zeste 2 polycomb repressive complex 2 subunit antibody
    • Enhancer of zeste homolog 2 (Drosophila) antibody
    • Enhancer of zeste homolog 2 antibody
    • Enhancer of zeste, Drosophila, homolog 2 antibody
    • ENX 1 antibody
    • Enx 1h antibody
    • ENX-1 antibody
    • ENX1 antibody
    • Enx1h antibody
    • EZH 2 antibody
    • EZH1 antibody
    • EZH2 antibody
    • EZH2_HUMAN antibody
    • EZH2b antibody
    • Histone-lysine N-methyltransferase EZH2 antibody
    • KMT 6 antibody
    • KMT6 antibody
    • KMT6A antibody
    • Lysine N-methyltransferase 6 antibody
    • MGC9169 antibody
    • WVS antibody
    • WVS2 antibody
    see all

Images

  • Chromatin was prepared from MDA-MB-231 cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 5µg of ab191250 (blue), and 20µl of Anti rabbit IgG sepharose beads. 5μg of rabbit normal IgG was added to the beads control (yellow).  The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).

    ChIP was performed according to the literature (PMID: 21880597).

  • All lanes : Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade (ab191250) at 1/1000 dilution

    Lane 1 : Wild-type HAP1 cell lysate
    Lane 2 : KMT6 / EZH2 knockout HAP1 cell lysate
    Lane 3 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
    Lane 4 : A549 (Human lung carcinoma cell line) whole cell lysate

    Lysates/proteins at 20 µg per lane.


    Predicted band size : 85 kDa
    Observed band size : 90 kDa (why is the actual band size different from the predicted?)

    Merged signal (red and green). Green - ab191250 observed at 93 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab191250 was shown to specifically recognize KMT6/EZH2 in wild-type HAP1 cells along with additional cross-reactive bands. No band was observed when KMT6/EZH2 knockout samples were examined. Wild-type and KMT6/EZH2 knockout samples were subjected to SDS-PAGE. ab191250 and ab8245 (loading control to GAPDH) were diluted at 1/1000 and 1/10,000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

  • All lanes : Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade (ab191250) at 1/5000 dilution

    Lane 1 : MCF7 (Human breast adenocarcinoma cell line) whole cell lysate
    Lane 2 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate

    Lysates/proteins at 20 µg per lane.

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

    Predicted band size : 85 kDa
    Observed band size : 90 kDa (why is the actual band size different from the predicted?)


    Exposure time : 30 seconds

    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade (ab191250) at 1/1000 dilution + U-2 OS (Human bone osteosarcoma epithelial cell line) whole cell lysate at 20 µg

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

    Predicted band size : 85 kDa
    Observed band size : 90 kDa (why is the actual band size different from the predicted?)


    Exposure time : 30 seconds

    Blocking/Dilution buffer: 5% NFDM/TBST.

  • KMT6 / EZH2 was immunoprecipitated from 0.35 mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab191250 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab191250 at 1/1000 dilution. VeriBlot for IP secondary antibody (HRP) (ab131366), was used as secondary antibody at 1/10000 dilution.

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

    Lane 2: ab191250 IP in HeLa whole cell lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab191250 in HeLa whole cell lysate.

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

    Exposure time: 3 minutes.

References

ab191250 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

Application
ChIP
Sample
Human Cell lysate - nuclear (Human B cells)
Negative control
Isotype IgG
Specification
Human B cells
Detection step
Real-time PCR
Type
Cross-linking (X-ChIP)
Duration of cross-linking step: 5 minute(s) and 0 second(s)
Specification of the cross-linking agent: Formaldehyde
Positive control
Gene specific Promoter primers
Username

Abcam user community

Verified customer

Submitted Jul 05 2017

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

Sign up