Anti-Filamin A antibody [EP2405Y]

5 product images

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  Western blot - Anti-Filamin A antibody [EP2405Y] (ab76289)

  Lane 1: Wild type HAP1 whole cell lysate (20 μg)
  Lane 2: FLNA knockout HAP1 whole cell lysate (20 μg)
  Lane 3: HepG2 whole cell lysate (20 μg)
  Lane 4: HeLa whole cell lysate (20 μg)
  

  Lanes 1 - 4: Merged signal (red and green). Green - ab76289 observed at 281 kDa. Red - loading control, ab18058, observed at 130 kDa.

  ab76289 was shown to specifically react with FLNA when FLNA knockout samples were used. Wild-type and FLNA knockout samples were subjected to SDS-PAGE. ab76289 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 250000 dilution and 1/10000 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 1/10000 dilution for 1 hour at room temperature before imaging.

  All lanes: Western blot - Anti-Filamin A antibody [EP2405Y] (ab76289)

  Predicted band size: 281 kDa

• 

  Western blot - Anti-Filamin A antibody [EP2405Y] (ab76289)

  Blocking buffer: 5% NFDM/TBST
  

  Diluting buffer: 5% NFDM/TBST

  All lanes: Western blot - Anti-Filamin A antibody [EP2405Y] (ab76289) at 1/10000 dilution

  Lane 1: HeLa (human cervix adenocarcinoma) whole cell lysate at 10 µg

  Lane 2: COS-1 (Cercopithecus aethiops kidney) whole cell lysate at 10 µg

  Secondary

  All lanes: Goat Anti-Rabbit IgG, (H+L), HRP conjugated at 1/1000 dilution

  Predicted band size: 281 kDa

  Observed band size: 281 kDa

  Exposure time: 30s

• 

  Western blot - Anti-Filamin A antibody [EP2405Y] (ab76289)

  All lanes: Western blot - Anti-Filamin A antibody [EP2405Y] (ab76289) at 1/500000 dilution

  Lane 1: COS-1 (African green monkey kidney fibroblast-like cell line) cell lysate at 10 µg

  Lane 2: Hela cell lysate at 10 µg

  Lane 3: 3T3 cell lysate at 10 µg

  Lane 4: C6 cell lysate at 10 µg

  Secondary

  All lanes: HRP labelled goat anti-rabbit at 1/1000 dilution

  Predicted band size: 281 kDa

  Observed band size: 281 kDa

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  Flow Cytometry (Intracellular) - Anti-Filamin A antibody [EP2405Y] (ab76289)

  Overlay histogram showing A431 cells stained with ab76289 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton 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 (ab76289 , 1/100 dilutio) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (Goat Anti-Rabbit IgG H&L (DyLight® 488) preadsorbed ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1μg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in A431 cells fixed with 4% paraformaldehyde/permeabilized in 0.1% PBS-Triton used under the same conditions.

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  Flow Cytometry (Intracellular) - Anti-Filamin A antibody [EP2405Y] (ab76289)

  ab76289 staining Filamin Ain the human cell line HeLa (human cervix adenocarcinoma) by intracellular flow cytometry. Cells were fixed with 4% paraformaldehyde, permeabilized with 90% methanol and the sample was incubated with the primary antibody at a dilution of 1/20. A goat anti rabbit IgG (Alexa Fluor^® 488) at a dilution of 1/2000 was used as the secondary antibody.
  

  Isoytype control: Rabbit monoclonal IgG (Black)

  Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue)