Anti-Amino-terminal enhancer of split/AES antibody [EPR8385]

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Amino-terminal enhancer of split/AES antibody [EPR8385] (AB137060)

Immunofluorescent staining of HeLa cells labelling Amino-terminal enhancer of split/AES using ab137060 at 1/250 dilution.

• WB

Lab

Western blot - Anti-Amino-terminal enhancer of split/AES antibody [EPR8385] (AB137060)

Lanes 1-4 : Merged signal (red and green). Green - ab137060 observed at 21 kDa. Red - loading control ab8245 observed at 36 kDa.

ab137060 Anti-Amino-terminal enhancer of split/AES antibody [EPR8385] was shown to specifically react with Amino-terminal enhancer of split/AES in wild-type HCT cells. Loss of signal was observed when knockout cell line ab266886 (knockout cell lysate ab257818) was used. Wild-type and Amino-terminal enhancer of split/AES knockout samples were subjected to SDS-PAGE. ab137060 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Amino-terminal enhancer of split/AES antibody [EPR8385] (ab137060) at 1/1000 dilution

Lane 1:

Wild-type HCT 116 (Human colorectal carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

AES knockout HCT 116 (Human colorectal carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

Western blot - Human AES (Amino-terminal enhancer of split) knockout HCT116 cell line (<a href='/en-us/products/cell-lines/human-aes-amino-terminal-enhancer-of-split-knockout-hct116-cell-line-ab266886'>ab266886</a>)

Lane 3:

HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg

Lane 4:

PC3 (Human prostate adenocarcinoma cell line) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 21 kDa

Observed band size: 21 kDa

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• WB

Lab

Western blot - Anti-Amino-terminal enhancer of split/AES antibody [EPR8385] (AB137060)

Western blot : Anti-Amino-terminal enhancer of split/AES antibody [EPR8385] ab137060 staining at 1/1000 dilution, shown in green; Mouse anti alpha Tubulin (ab7291) loading control staining at 1/20,000 dilution, shown in magenta. A band was observed at 24 kDa in Wild-type A549 cell lysates with no signal observed at this size in TLE5 knockout A549 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5pc 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 800CW & Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Amino-terminal enhancer of split/AES antibody [EPR8385] (ab137060) at 1/1000 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

TLE5 knockout A549 at 20 µg

Lane 3:

HeLa at 20 µg

Lane 4:

THP-1 at 20 µg

Secondary

Lanes 1 - 4:

Goat anti-Rabbit 800CW at 1/20000 dilution

Lanes 1 - 4:

Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 24 kDa

Observed band size: 24 kDa,51 kDa

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• WB

Unknown

Western blot - Anti-Amino-terminal enhancer of split/AES antibody [EPR8385] (AB137060)

All lanes:

Western blot - Anti-Amino-terminal enhancer of split/AES antibody [EPR8385] (ab137060) at 1/1000 dilution

Lane 1:

293T cell lysate at 10 µg

Lane 2:

HeLa cell lysate at 10 µg

Lane 3:

PC3 cell lysate at 10 µg

Lane 4:

Jurkat cell lysate at 10 µg

Secondary

All lanes:

HRP labelled Goat anti-Rabbit IgG at 1/2000 dilution

Predicted band size: 21 kDa

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