Anti-Cyclin E2 antibody [EP454Y] - BSA and Azide free

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Cyclin E2 antibody [EP454Y] - BSA and Azide free (AB239833)

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 ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1 : 200 (2.5 µg/ml). Goat anti rabbit IgG (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.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (anti cyclin e2 antibody ep454y immunocytochemistry hela human)

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cyclin E2 antibody [EP454Y] - BSA and Azide free (AB239833)

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

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab40890).

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cyclin E2 antibody [EP454Y] - BSA and Azide free (AB239833)

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 using ab93684 (Tris/EDTA buffer, pH 9.0). 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. This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab40890)

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Cyclin E2 antibody [EP454Y] - BSA and Azide free (AB239833)

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 Fluor^® 488) (ab150077) was used as the secondary antibody at a 1/2000 dilution.

Left panel - Rabbit monoclonal IgG (ab172730).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab40890).

• IP

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Immunoprecipitation - Anti-Cyclin E2 antibody [EP454Y] - BSA and Azide free (AB239833)

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 (ab172730) instead of ab40890 in Jurkat whole cell lysate
For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/5000 dilution.
Blocking and diluting buffer : 5% NFDM/TBST. This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab40890)

All lanes:

Immunoprecipitation - Anti-Cyclin E2 antibody [EP454Y] (<a href='/en-us/products/primary-antibodies/cyclin-e2-antibody-ep454y-ab40890'>ab40890</a>)

Predicted band size: 47 kDa

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• WB

Lab

Western blot - Anti-Cyclin E2 antibody [EP454Y] - BSA and Azide free (AB239833)

This data was developed using the same antibody clone in a different buffer formulation (ab40890).

Lanes 1 - 4 : Merged signal (red and green). Green - ab40890 observed at 45 kDa. Red - 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 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 (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 800CW) preabsorbed (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] (<a href='/en-us/products/primary-antibodies/cyclin-e2-antibody-ep454y-ab40890'>ab40890</a>) 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

Predicted band size: 47 kDa

Observed band size: 45 kDa

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