Anti-Zyxin antibody [ZOL301]

• WB

Lab

Western blot - Anti-Zyxin antibody [ZOL301] (AB50391)

Lanes 1 - 4 : Merged signal (red and green). Green - ab50391 observed at 75 kDa. Red - loading control ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) observed at 37 kDa.

ab50391 was shown to react with ZYX in wild-type HEK-293T cells in Western blot with loss of signal observed in ZYX knockout cell line ab266504 (ZYX knockout cell lysate ab257810). Wild-type HEK-293T and ZYX knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween^®) before incubation with ab50391 and ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye^® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-Zyxin antibody [ZOL301] (ab50391) at 1/1000 dilution

Lane 1:

Wild-type HEK-293T cell lysate at 20 µg

Lane 2:

ZYX knockout HEK-293T cell lysate at 20 µg

Lane 2:

Western blot - Human ZYX (Zyxin) knockout HEK-293T cell line (<a href='/en-us/products/cell-lines/human-zyx-zyxin-knockout-hek-293t-cell-line-ab266504'>ab266504</a>)

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

Daudi cell lysate at 20 µg

Predicted band size: 61 kDa

Observed band size: 75 kDa

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• WB

Lab

Western blot - Anti-Zyxin antibody [ZOL301] (AB50391)

False colour image of Western blot : Anti-Zyxin antibody [ZOL301] staining at 1/1000 dilution, shown in green; Rabbit Anti-GAPDH antibody [EPR16891] (ab181602) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab50391 was shown to bind specifically to Zyxin. A band was observed at 75 kDa in wild-type HEK-293T cell lysates with no signal observed at this size in ZYX CRISPR-Cas9 edited cell line ab266503 (CRISPR-Cas9 edited cell lysate ab257809). The band observed in the CRISPR-Cas9 edited lysate lane below 75 kDa is likely to represent a truncated form of Zyxin. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and ZYX CRISPR-Cas9 edited HEK-293T cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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-Mouse IgG H&L (IRDye^® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preabsorbed (ab216777) at 1/20000 dilution.

All lanes:

Western blot - Anti-Zyxin antibody [ZOL301] (ab50391) at 1/1000 dilution

Lane 1:

Wild-type HEK-293T cell lysate at 20 µg

Lane 2:

ZYX CRISPR-Cas9 edited HEK-293T cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

Daudi cell lysate at 20 µg

Predicted band size: 61 kDa

Observed band size: 75 kDa

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