Anti-HLA A antibody [EP1395Y] (ab52922)

Anti-HLA A antibody [EP1395Y] images

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  IHC - Wholemount - Anti-HLA A [EP1395Y] antibody (ab52922)This image is courtesy of an Abreview submitted by Rita Fior

  IHC - Wholemount of human zebrafish xenograft labelling HLA A with ab52922. Sample was incubated with primary antibody (1/100) for 1 hours at 4°C. An Alexa Fluor^® 647-conjugated goat anti-rabbit IgG polyclonal (1/400) was used as the secondary antibody.

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  Immunocytochemistry/ Immunofluorescence - Anti-HLA A antibody [EP1395Y] (ab52922)

  ICC/IF image of unpurified ab52922 stained MCF7 cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab52922, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
  
  

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  Western blot - Anti-HLA A antibody [EP1395Y] (ab52922)
  All lanes : Anti-HLA A antibody [EP1395Y] (ab52922) at 1/5000 dilution (purified)
  
  Lane 1 : THP-1 cell lysate at 20 µg
  Lane 2 : A549 cell lysate at 1/20 dilution
  
  Secondary
  Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)
  
  Predicted band size : 41 kDa
  

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

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  Western blot - Anti-HLA A antibody [EP1395Y] (ab52922)
  Anti-HLA A antibody [EP1395Y] (ab52922) at 1/2000 dilution (purified) + HL-60 cell lysate at 20 µg
  
  Secondary
  Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/1000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)
  
  Predicted band size : 41 kDa
  

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

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  Western blot - Anti-HLA A [EP1395Y] antibody (ab52922)
  Anti-HLA A antibody [EP1395Y] (ab52922) at 1/10000 dilution + Raji cell lysate at 10 µg
  
  Secondary
  Goat anti rabbit IgG HRP conjugated at 1/2000 dilution
  
  Predicted band size : 41 kDa
  Observed band size : 41 kDa
  
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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HLA A antibody [EP1395Y] (ab52922)
  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human tonsil tissue labelling HLA A with purified ab52922 at 1/100. Heat mediated antigen retrieval was performed using EDTA buffer pH 9. ab97051, a goat anti-rabbit IgG H&L (HRP) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HLA A antibody [EP1395Y] (ab52922)This image is courtesy of an anonymous Abreview

  ab52922 staining HLA A in Human tonsil tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 3% H₂O₂ for 10 minutes at 25°C; antigen retrieval was by heat mediation in a citrate buffer, pH 6.0 . Samples were incubated with primary antibody (1/3000) for 20 minutes at 25°C. An undiluted HRP-conjugated Goat anti-rabbit IgG polyclonal was used as the secondary antibody.

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HLA A antibody [EP1395Y] (ab52922)

  Ab52922 at 1/250 dilution staining human tonsil; paraffin embedded.

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  Immunocytochemistry/ Immunofluorescence - Anti-HLA A antibody [EP1395Y] (ab52922)

  Immunocytochemistry/Immunofluorescence analysis of Raji (human Burkitt's lymphoma) cells labelling HLA A with purified ab52922 at 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/1000) were also used.

  Control 1: primary antibody (1/100) and secondary antibody, ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/500).

  Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/500).

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  Flow Cytometry - Anti-HLA A antibody [EP1395Y] (ab52922)

  Flow Cytometry analysis of Raji cells labelling HLA A with purified ab52922 at 1/40 (red). Cells were fixed with 2% paraformaldehyde. A FITC-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

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  Flow Cytometry - Anti-HLA A [EP1395Y] antibody (ab52922)
  Overlay histogram showing Raji cells stained with ab52922 (red line). The cells were fixed with 4% paraformaldehyde (10 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 (ab52922, 1/100) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. 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 Raji cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
  
  
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  Immunoprecipitation - Anti-HLA A antibody [EP1395Y] (ab52922)

  ab52922 (purified) at 1/20 immunoprecipitating HLA A in THP-1 whole cell lysate. 10 ug of cell lysate was present in the input. For western blotting, a HRP-conjugated Veriblot for IP secondary antibody (ab131366) (1/10,000) was used as the secondary antibody. A rabbit monoclonal IgG (ab172730) was used intead of ab128913 as a negative control (Lane 3).

  Blocking buffer and concentration: 5% NFDM/TBST.

  Diluting buffer and concentration: 5% NFDM /TBST.

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  Immunoprecipitation - Anti-HLA A antibody [EP1395Y] (ab52922)

  ab52922 (purified) at 1/20 immunoprecipitating HLA A in A549 whole cell lysate. 10 ug of cell lysate was present in the input. For western blotting, a HRP-conjugated Veriblot for IP secondary antibody (ab131366) (1/10,000) was used as the secondary antibody. A rabbit monoclonal IgG (ab172730) was used intead of ab128913 as a negative control (Lane 3).

  Blocking buffer and concentration: 5% NFDM/TBST.

  Diluting buffer and concentration: 5% NFDM /TBST.