Anti-KMT6 / EZH2 antibody [EPR20108] - BSA and Azide free

3 product images

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  Western blot - Anti-KMT6 / EZH2 antibody [EPR20108] - BSA and Azide free (ab240992)

  Merged signal (red and green). Green - Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250 observed at 93 kDa. Red - loading control, Anti-GAPDH antibody [6C5] - Loading Control ab8245, observed at 37 kDa.
  

  Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade 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. Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250 and Anti-GAPDH antibody [6C5] - Loading Control 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 (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (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.

  This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250).

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

  Lane 1: Wild-type HAP1 cell lysate at 20 µg

  Lane 2: KMT6 / EZH2 knockout HAP1 cell lysate at 20 µg

  Lane 3: HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg

  Lane 4: MCF7 (human breast adenocarcinoma cell line) whole cell lysate at 20 µg

  Predicted band size: 85 kDa

  Observed band size: 90 kDa

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  Immunoprecipitation - Anti-KMT6 / EZH2 antibody [EPR20108] - BSA and Azide free (ab240992)

  KMT6 / EZH2 was immunoprecipitated from 0.35 mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250 at 1/30 dilution. Western blot was performed from the immunoprecipitate using Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366), was used for detection at 1/10000 dilution.
  

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

  Lane 2: Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250 IP in HeLa whole cell lysate.

  Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250 in HeLa whole cell lysate.

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

  Exposure time: 3 minutes.

  This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250).

  All lanes: Immunoprecipitation - Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade (Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250)

  Predicted band size: 85 kDa

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  Western blot - Anti-KMT6 / EZH2 antibody [EPR20108] - BSA and Azide free (ab240992)

  False colour image of Western blot: Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-KMT6 / EZH2 antibody [EPR20108] - ChIP Grade ab191250 was shown to bind specifically to KMT6 / EZH2. A band was observed at 90 kDa in wild-type MCF7 cell lysates with no signal observed at this size in ezh2 CRISPR-Cas9 edited cell line ab281611 (CRISPR-Cas9 edited cell lysate ab282963). The band observed in the CRISPR-Cas9 edited lysate lane below 90 kDa is likely to represent a truncated form of KMT6 / EZH2. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and ezh2 CRISPR-Cas9 edited MCF7 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

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

  Lane 1: Wild-type MCF7 cell lysate at 20 µg

  Lane 2: ezh2 CRISPR-Cas9 edited MCF7 cell lysate at 20 µg

  Lane 3: HEK-293 cell lysate at 20 µg

  Lane 4: Mouse Testis cell lysate at 20 µg

  Performed under reducing conditions.

  Observed band size: 90 kDa