HRP Anti-Smad4 antibody [EP618Y]

• WB

Lab

Western blot - HRP Anti-Smad4 antibody [EP618Y] (AB195554)

This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 3% milk before being incubated with ab195554 overnight at 4°C. Antibody binding was visualised using ECL development solution ab133406.

All lanes:

Western blot - HRP Anti-Smad4 antibody [EP618Y] (ab195554) at 1/5000 dilution

All lanes:

SHSY-5Y (Human neuroblastoma cell line) Whole Cell Lysate at 10 µg

Predicted band size: 60 kDa,82 kDa

Observed band size: 30 kDa,47 kDa,52 kDa,54 kDa,65 kDa,82 kDa

true

Exposure time: 20min

• WB

Lab

Western blot - HRP Anti-Smad4 antibody [EP618Y] (AB195554)

ab195554 was shown to recognize Smad4 in wild-type HAP1 cells as signal was lost at the expected MW in SMAD4 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and SMAD4 knockout samples were subjected to SDS-PAGE. ab195554 and ab184095 (Mouse monoclonal [mAbcam 9484] to GAPDH - Loading Control (Alexa Fluor^® 680) loading control) were incubated overnight at 4°C at 1/5000 dilution and 1/20000 dilution respectively. The loading control was imaged using the Licor Odyssey CLx prior to blots being developed with ECL technique.

All lanes:

Western blot - HRP Anti-Smad4 antibody [EP618Y] (ab195554) at 1/5000 dilution

Lane 1:

Wild-type HAP1 whole cell lysate at 20 µg

Lane 2:

SMAD4 knockout HAP1 whole cell lysate at 20 µg

Predicted band size: 60 kDa

Observed band size: 65 kDa

false

Exposure time: 12min