Anti-FTO antibody [EPR6894]

8 product images

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  Western blot - Anti-FTO antibody [EPR6894] (ab126605)

  Lane 1: Wild-type HAP1 whole cell lysate (20 μg)
  Lane 2: FTO knockout HAP1 whole cell lysate (20 μg)
  Lane 3: HEK293 whole cell lysate (20 μg)
  Lane 4: MOLT4 whole cell lysate (20 μg)
  

  Lanes 1 - 4: Merged signal (red and green). Green - ab126605 observed at 58 kDa. Red - loading control, Anti-GAPDH antibody [6C5] - Loading Control ab8245, observed at 37 kDa.

  ab126605 was shown to specifically react with FTO in wild-type HAP1 cells. No band was observed when FTO knockout samples were examined. Wild-type and FTO knockout samples were subjected to SDS-PAGE. ab126605 and Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at a 1/1000 dilution and 1/10,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (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: Western blot - Anti-FTO antibody [EPR6894] (ab126605)

  Predicted band size: 58 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FTO antibody [EPR6894] (ab126605)

  Immunohistochemical staining of paraffin embedded human hepatocellular carcinoma with purified ab126605 at a working dilution of 1 in 500. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

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  Immunocytochemistry/ Immunofluorescence - Anti-FTO antibody [EPR6894] (ab126605)

  Immunofluorescence staining of BxPC-3 cells with purified ab126605 at a working dilution of 1 in 250, counter-stained with DAPI. The secondary antibody was Alexa Fluor^® 555 goat anti rabbit, used at a dilution of 1 in 500. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative control is shown in bottom right hand panel - for the negative control, purified ab126605 was used at a dilution of 1/200 followed by an Alexa Fluor^® 488 goat anti-mouse antibody at a dilution of 1/500.

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  Western blot - Anti-FTO antibody [EPR6894] (ab126605)

  Blocking buffer: 5% NFDM/TBST

  Dilution buffer: 5% NFDM/TBST

  All lanes: Western blot - Anti-FTO antibody [EPR6894] (ab126605) at 1/10000 dilution

  Lane 1: Molt-4 cell lysate at 20 µg

  Lane 2: HEK293 cell lysate at 20 µg

  Lane 3: BxPC-3 cell lysate at 20 µg

  Lane 4: Caco-2 cell lysate at 20 µg

  Lane 5: SH-SY5Y cell lysate at 20 µg

  Secondary

  All lanes: HRP goat anti-rabbit (H+L) at 1/1000 dilution

  Predicted band size: 58 kDa

  Observed band size: 58 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-FTO antibody [EPR6894] (ab126605)

  Immunohistochemical staining of FTO in paraffin embedded human adrenal gland tissue using unpurified ab126605 at a 1/50 dilution.

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  Immunocytochemistry/ Immunofluorescence - Anti-FTO antibody [EPR6894] (ab126605)

  Unpurified ab126605 at a 1/50 dilution staining FTO in BxPC3 cells by immunofluorescence.

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  Western blot - Anti-FTO antibody [EPR6894] (ab126605)

  All lanes: Western blot - Anti-FTO antibody [EPR6894] (ab126605) at 1/1000 dilution

  Lane 1: MOLT4 cell lysate at 10 µg

  Lane 2: 293T cell lysate at 10 µg

  Lane 3: SH SY5Y cell lysate at 10 µg

  Lane 4: BxPC3 cell lysate at 10 µg

  Lane 5: Caco 2 cell lysate at 10 µg

  Secondary

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

  Predicted band size: 58 kDa

  Observed band size: 58 kDa

• 

  Western blot - Anti-FTO antibody [EPR6894] (ab126605)

  False colour image of Western blot: Anti-FTO antibody [EPR6894] 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, ab126605 was shown to bind specifically to FTO. A band was observed at 58 kDa in wild-type MCF7 cell lysates with no signal observed at this size in FTO knockout cell line. To generate this image, wild-type and FTO knockout 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-FTO antibody [EPR6894] (ab126605) at 1/1000 dilution

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

  Lane 2: Western blot - Human FTO knockout MCF7 cell line (Human FTO knockout MCF7 cell line ab282631)

  Lane 2: FTO knockout MCF7 cell lysate at 20 µg

  Lane 3: HEK-293 cell lysate at 20 µg

  Lane 4: MOLT-4 cell lysate at 20 µg

  Secondary

  All lanes: Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

  Performed under reducing conditions.

  Predicted band size: 58 kDa

  Observed band size: 58 kDa