Anti-DEPDC5 antibody [EPR20497-23] - BSA and Azide free

• WB

Lab

Western blot - Anti-DEPDC5 antibody [EPR20497-23] - BSA and Azide free (AB272399)

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

The wild-type and DEPDC5 knockout mouse E14 brain tissue lysates were kindly provided by an anonymous collaborator.

ab213181 was shown to specifically react with DEPDC5 in wild-type mouse E14 brain tissue as signal was lost in DEPDC5 knockout tissue. Wild-type and DEPDC5 knockout samples were subjected to SDS-PAGE. ab213181 and ab129002 (Rabbit anti-Vinculin loading control) were incubated 1 hour at room temperature at 1/1000 dilution and 1/5000 dilution respectively. Blots were developed with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) secondary antibody at 1/100, 000 dilution for 1 hour at room temperature before imaging. The blot was developed on a BIO-RAD^® ChemiDoc™ MP instrument using the ECL technique.

Exposure time : 26 seconds.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, glycerol, BSA and sodium azide (ab213181).

All lanes:

Western blot - Anti-DEPDC5 antibody [EPR20497-23] (<a href='/en-us/products/primary-antibodies/depdc5-antibody-epr20497-23-ab213181'>ab213181</a>) at 1/1000 dilution

Lane 1:

Wild-type mouse E14 brain tissue lysate at 20 µg

Lane 2:

DEPDC5 knockout mouse E14 brain tissue lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 178 kDa

Observed band size: 181 kDa

false

• WB

Lab

Western blot - Anti-DEPDC5 antibody [EPR20497-23] - BSA and Azide free (AB272399)

False colour image of Western blot : Anti-DEPDC5 antibody [EPR20497-23] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab213181 was shown to bind specifically to DEPDC5. A band was observed at 181 kDa in wild-type A549 cell lysates with no signal observed at this size in DEPDC5 knockout cell line ab266906 (knockout cell lysate ab258394). To generate this image, wild-type and DEPDC5 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged.Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-DEPDC5 antibody [EPR20497-23] (<a href='/en-us/products/primary-antibodies/depdc5-antibody-epr20497-23-ab213181'>ab213181</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

DEPDC5 knockout A549 cell lysate at 20 µg

Lane 2:

Western blot - Human DEPDC5 knockout A549 cell line (<a href='/en-us/products/cell-lines/human-depdc5-knockout-a549-cell-line-ab266906'>ab266906</a>)

Lane 3:

Human brain tissue lysate at 20 µg

Lane 4:

Mouse brain tissue lysate at 20 µg

Predicted band size: 178 kDa

Observed band size: 181 kDa

false