Anti-Cyclin E2 antibody [EP454Y]

7 product images

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  Western blot - Anti-Cyclin E2 antibody [EP454Y] (ab40890)

  Lanes 1 - 4: Merged signal (red and green). Green - ab40890 observed at 45 kDa. Red - loading control, Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.
  

  ab40890 was shown to react with CCNE2 in HAP1 wild-type cells in Western blot. Loss of signal was observed when CCNE2 knockout sample was used. HAP1 wild-type and CCNE2 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% Milk in TBS-T (0.1% Tween^®) before incubation with ab40890 and Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 10000 dilution and a 1 in 20000 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^® 800CW) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

  All lanes: Western blot - Anti-Cyclin E2 antibody [EP454Y] (ab40890) at 1/10000 dilution

  Lane 1: Wild-type HAP1 whole cell lysate at 20 µg

  Lane 2: CCNE2 knockout HAP1 whole cell lysate at 20 µg

  Lane 3: MCF7 whole cell lysate at 20 µg

  Lane 4: HeLa whole cell lysate at 20 µg

  Performed under reducing conditions.

  Predicted band size: 47 kDa

  Observed band size: 45 kDa

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  Immunoprecipitation - Anti-Cyclin E2 antibody [EP454Y] (ab40890)

  ab40890 (purified) at 1/20 dilution (1 µg) immunoprecipitating Cyclin E2 in Jurkat whole cell lysate.
  Lane 1 (input): Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10 µg
  Lane 2 (+): ab40890 & Jurkat whole cell lysate
  Lane 3 (-): Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab40890 in Jurkat whole cell lysate
  For western blotting, VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) was used at 1/5000 dilution.
  Blocking and diluting buffer: 5% NFDM/TBST.

  All lanes: Immunoprecipitation - Anti-Cyclin E2 antibody [EP454Y] (ab40890)

  Predicted band size: 47 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cyclin E2 antibody [EP454Y] (ab40890)

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast carcinoma tissue sections labeling Cyclin E2 with purified ab40890 at 1/100 dilution (1.41 µg/ml). Heat mediated antigen retrieval was performed Perform heat mediated antigen retrieval using Antigen Retrieval Buffer (100X Tris-EDTA Buffer, pH 9.0) ab93684 (Tris/EDTA buffer, pH 9.0). Rabbit specific IHC polymer detection kit HRP/DAB (Rabbit specific IHC polymer detection kit HRP/DAB ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

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  Immunocytochemistry/ Immunofluorescence - Anti-Cyclin E2 antibody [EP454Y] (ab40890)

  Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Cyclin E2 with purified ab40890 at 1:50 dilution (2.8 μg/ml). Cells were fixed in 100% Methanol and permeabilized with None. Cells were counterstained with Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1:200 (2.5 μg/ml). Goat anti rabbit IgG (Alexa Fluor^® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) was used as the secondary antibody at 1:1000 (2 μg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

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  Western blot - Anti-Cyclin E2 antibody [EP454Y] (ab40890)

  All lanes: Western blot - Anti-Cyclin E2 antibody [EP454Y] (ab40890) at 1/1000 dilution

  All lanes: Jurkat (Human T cell leukemia T lymphocyte) whole cell lysates at 15 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: 47 kDa

  Observed band size: 45 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cyclin E2 antibody [EP454Y] (ab40890)

  ab40890 (unpurified) at 1/250 staining human breast carcinoma by immunohistochemistry, paraffin-embedded tissue.

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  Flow Cytometry (Intracellular) - Anti-Cyclin E2 antibody [EP454Y] (ab40890)

  Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling Cyclin E2 (right) with purified ab40890 at a 1/20 dilution. Cells were fixed with 90% ethanol and permeabilized with 90% methanol. A goat anti-rabbit IgG (Alexa Fluorr^® 488) (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) was used as the secondary antibody at a 1/2000 dilution.
  

  Left panel - Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730).