Anti-IMP3 antibody [EPR12021]

6 product images

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  Western blot - Anti-IMP3 antibody [EPR12021] (ab177477)

  Lane 1: Wild-type HAP1 whole cell lysate (20 μg)
  Lane 2: IGF2BP3 (IMP3) 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 - ab177477 observed at 70 kDa. Red - loading control, Anti-GAPDH antibody [mAbcam 9484] - Loading Control ab9484, observed at 37 kDa.

  ab177477 was shown to specifically react with IGF2BP3 (IMP3) in wild-type HAP1 cells along with additional cross-reactive bands. No band was observed when IGF2BP3 (IMP3) knockout cells were examined. Wild-type and IGF2BP3 (IMP3) knockout samples were subjected to SDS-PAGE. ab177477 and Anti-GAPDH antibody [mAbcam 9484] - Loading Control ab9484 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20,000 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 1/20,000 dilution for 1 hour at room temperature before imaging.

  All lanes: Western blot - Anti-IMP3 antibody [EPR12021] (ab177477)

  Predicted band size: 63 kDa

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  Flow Cytometry (Intracellular) - Anti-IMP3 antibody [EPR12021] (ab177477)

  Intracellular Flow Cytometry analysis of BxPC-3 (human pancreas adenocarcinoma) cells labeling with purified ab177477 at 1/300 dilution (10ug/ml) (Red). Cells were fixed with 80% methanoland permeabilised with 0.1% Tween-20. A Goat anti rabbit IgG (Alexa Fluor^® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) was used as a isotype control. Cell without incubation with primary antibody and secondary antibody (Blue) were used as unlabeled control.

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  Western blot - Anti-IMP3 antibody [EPR12021] (ab177477)

  All lanes: Western blot - Anti-IMP3 antibody [EPR12021] (ab177477) at 1/10000 dilution

  Lane 1: MDA-MB-231 (Human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 2: MDA-MB-435S (Human mammary gland ductal carcinoma melanocyte) whole cell lysate at 20 µg

  Lane 3: HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

  Lane 4: HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate at 20 µg

  Lane 5: Mouse placenta lysate at 20 µg

  Secondary

  All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

  Predicted band size: 63 kDa

  Observed band size: 69 kDa

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  Immunoprecipitation - Anti-IMP3 antibody [EPR12021] (ab177477)

  Immunoprecipitation of IMP3 from 293T (Human embryonic kidney epithelial cell) whole cell lysate using ab177477 at 1/20 dilution.
  
  Lane 1: 293T (Human embryonic kidney epithelial cell) whole cell lysate 10μg
  
  Lane 2: 293T whole cell lysate
  
  Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab177477 in 293T whole cell lysate
  
  VeriBlot for IP Detection Reagent (HRP) ab131366 was used as secondary antibody at 1/1000 dilution.
  
  Blocking and diluting buffer and concentration: 5% NFDM/TBST.
  
  Observed MW at 69kDa.

  All lanes: Immunoprecipitation - Anti-IMP3 antibody [EPR12021] (ab177477) at 1/20 dilution

  Lane 1: 293T (Human embryonic kidney epithelial cell) whole cell lysate at 10 µg

  Lane 2: 293T whole cell lysate

  Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab177477 in 293T whole cell lysate

  Secondary

  All lanes: Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) at 1/1000 dilution

  Predicted band size: 63 kDa

  Observed band size: 69 kDa

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  Western blot - Anti-IMP3 antibody [EPR12021] (ab177477)

  Blocking and diluting buffer and concentration: 5% NFDM/TBST.
  
  We are unsure about the nature of the extra bands.

  All lanes: Western blot - Anti-IMP3 antibody [EPR12021] (ab177477) at 1/10000 dilution

  Lane 1: Mouse spleen lysate at 20 µg

  Lane 2: Rat heart lysate at 20 µg

  Lane 3: Rat spleen lysate at 20 µg

  Secondary

  All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

  Predicted band size: 63 kDa

  Observed band size: 69 kDa

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  Immunocytochemistry/ Immunofluorescence - Anti-IMP3 antibody [EPR12021] (ab177477)

  Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling IMP3 using ab177477. The cells were fixed with 4% paraformaldehyde then permeabilized with 0.1% Triton X-100. The cells were then incubated with ab177477 at 1:50 dilution followed by a further incubation with a Goat anti rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI. Cells were counterstained using Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1:200 dilution (shown in red). Secondary antibody only control: PBS instead of the primary antibody.