Anti-Optineurin antibody [EPR23059-124]

6 product images

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  Western blot - Anti-Optineurin antibody [EPR23059-124] (ab242146)

  Blocking and diluting buffer and concentration: 5% NFDM/TBST The molecular weight observed is consistent with what has been described in the literature (PMID: 27620379).
  

  Exposure time: Lane 1: 15 seconds Lane 2: 70 seconds

  All lanes: Western blot - Anti-Optineurin antibody [EPR23059-124] (ab242146) at 1/1000 dilution

  Lane 1: Human brain lysate at 20 µg

  Lane 2: Hela (human cervix adenocarcinoma epithelial cell) 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 1/100000 dilution

  Predicted band size: 66 kDa

  Observed band size: 74 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Optineurin antibody [EPR23059-124] (ab242146)

  Immunohistochemical analysis of paraffin-embedded Human cerebrum tissue labeling Optineurin with ab242146 at 1/500 dilution (0.98 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (Rabbit specific IHC polymer detection kit HRP/DAB ab209101). Cytoplasmic staining on human cerebrum. The section was incubated with ab242146 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND®RX instrument. Counterstained with Hematoxylin.
  

  Secondary antibody only control: Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (Rabbit specific IHC polymer detection kit HRP/DAB ab209101).

  Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.

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  Flow Cytometry (Intracellular) - Anti-Optineurin antibody [EPR23059-124] (ab242146)

  Intracellular flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (human cervix adenocarcinoma) cells labelling Optineurin with ab242146 at 1/50 (Red) compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730, Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor^®488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) at 1/2000 dilution was used as the secondary antibody.
  

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  Immunoprecipitation - Anti-Optineurin antibody [EPR23059-124] (ab242146)

  Immunoprecipitation of OPTN in U20S cells. Lysates were prepared and immunoprecipitation was performed using 1.0 µg of ab242146 pre-coupled to Protein A beads. Samples were washed and processed for western blot with OPTN Monoclonal Antibody at 1/10000
  
  This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

  Lane 1: Immunoprecipitation at 1/10000 dilution

  Lane 3: Immunoprecipitation - Anti-Optineurin antibody [EPR23059-124] (ab242146) at 1 µg

  All lanes: U20S cells

  Observed band size: 66 kDa

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  Immunocytochemistry/ Immunofluorescence - Anti-Optineurin antibody [EPR23059-124] (ab242146)

  ab242146 was shown to react with OPTN in wild-type U20S cells in immunocytochemistry with loss of signal observed in a OPTN knockout cell line. Wild-type and knockout cells were mixed and pelleted at a 1:1 ratio on coverslips. The cells were fixed with 4% paraformaldehyde (15 min) then permeabilized with 0.1% Triton X-100 (10min) and then blocked with 1x PBS, 0.01% Triton X-100, 5% BSA, 5% NGS. The cells were then incubated with ab242146 at dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat anti-rabbit secondary antibody to (Alexa Fluor® 555) at 0.5 ?g/ml. Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880).
  
  This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

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  Western blot - Anti-Optineurin antibody [EPR23059-124] (ab242146)

  ab242146 was shown to react with OPTN in wild-type A431 cells in Western blot with loss of signal observed in OPTN knockout cell line ab261906. Wild-type A431 and OPTN knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab242146 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with secondary antibodies at 0.2ug/mL before imaging.
  
  This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

  All lanes: Western blot - Anti-Optineurin antibody [EPR23059-124] (ab242146) at 1/1000 dilution

  Lane 1: Wild-type A431 lysate at 20 µg

  Lane 2: Western blot - Human DAG1 knockout A-431 cell line (ab261906) at 20 µg

  Observed band size: 66 kDa