Anti-ROCK2 + ROCK1 antibody [EP786Y]

8 product images

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  Immunoprecipitation - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171)

  ab45171 (purified) at 1/40 immunoprecipitating ROCK2 + ROCK1 in HeLa cell lysate. For western blotting, a peroxidase-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/1000).
  

  Blocking buffer and concentration: 5% NFDM/TBST.

  Diluting buffer and concentration: 5% NFDM /TBST.

  All lanes: Immunoprecipitation - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171)

  Observed band size: 158 kDa

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  Western blot - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171)

  Blocking buffer and concentration: 5% NFDM/TBST.

  Diluting buffer and concentration: 5% NFDM /TBST.

  All lanes: Western blot - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171) at 1/5000 dilution

  Lane 1: HeLa cell lysate - treated with Calyculin A at 20 µg

  Lane 2: HeLa cell lysate - treated with Camptothecin at 20 µg

  Lane 3: HeLa cell lysate at 20 µg

  Lane 4: Jurkat cell lysate at 20 µg

  Lane 5: Ramos cell lysate at 20 µg

  Secondary

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

  Observed band size: 158 kDa

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  Immunocytochemistry/ Immunofluorescence - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171)

  Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling ROCK2 + ROCK1 with purified ab45171 at 1/50. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 555) ab150078, an Alexa Fluor^® 555-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain.
  

  Control: primary antibody (1/50) and secondary antibody, Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) ab150113, an Alexa Fluor^® 488-conjugated goat anti-mouse IgG (1/500).

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  Western blot - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171)

  Loading control: Anti-6X His tag® antibody [EPR20547] (Anti-6X His tag® antibody [EPR20547] - ChIP Grade ab213204)
  

  Blocking buffer and concentration: 5% NFDM/TBST

  Exposure Times:

  Lane 1: 3 seconds

  Lane 2: 7 seconds

  Lanes 1 - 2: Western blot - Alexa Fluor® 488 Anti-ROCK2 + ROCK1 antibody [EP786Y] (Alexa Fluor® 488 Anti-ROCK2 + ROCK1 antibody [EP786Y] ab205432) at 1/1000 dilution

  Lanes 1 - 2: Western blot - PE Anti-ROCK2 + ROCK1 antibody [EP786Y] (PE Anti-ROCK2 + ROCK1 antibody [EP786Y] ab224972) at 1/1000 dilution

  Lanes 1 - 2: Western blot - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171) at 1/1000 dilution

  Lane 1: Recombinant Human ROCK1 protein (aa 1114 to 1354) (30 kDa)

  Lane 2: Recombinant Human ROCK2 protein (aa 1132 to 1388) (30 kDa)

  Secondary

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

  Observed band size: 30 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171)

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human adenocarcinoma of the colon tissue labelling ROCK2 + ROCK1 with purified ab45171 at 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. A prediluted HRP-polymer conjugated anti-rabbit IgG was used as the secondary antibody. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

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  Western blot - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171)

  Blocking buffer and concentration: 5% NFDM/TBST.

  Diluting buffer and concentration: 5% NFDM /TBST.

  All lanes: Western blot - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171) at 1/5000 dilution

  Lane 1: PC-12 cell lysate at 20 µg

  Lane 2: RAW264.7 cell lysate at 20 µg

  Secondary

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

  Observed band size: 158 kDa

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  Flow Cytometry (Intracellular) - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171)

  Overlay histogram showing HeLa cells stained with unpurified ab45171 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween 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 (ab45171, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluorr^® 488 goat anti-rabbit IgG (H+L) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.
  

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ROCK2 + ROCK1 antibody [EP786Y] (ab45171)

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human thyroid gland carcinoma tissue labelling ROCK2 + ROCK1 with unpurified ab45171.
  

  Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.