Lane 1: Wild type HAP1 whole cell lysate (20 µg)
Lane 2: IKB alpha knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: HepG2 whole cell lysate (20 µg)

Lanes 1 - 4: Merged signal (red and green). Green - ab12134 observed at 38 kDa. Red - loading control, ab176560, observed at 50 kDa.

ab12134 was shown to specifically react with IKB alpha in wild type cells as signal was lost in IKB alpha knockout cells. Wild-type and IKB alpha knockout samples were subjected to SDS-PAGE. Ab12134 and ab176560 (Rabbit anti alpha Tubulin loading control) were incubated overnight at 4°C at 1 ug/ml and 1/10000 dilution respectively. Blots were developed 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 1/10000 dilution for 1 hour at room temperature before imaging.

ab12134 detects both phosphorylated and non-phosphorylated forms of IKB alpha by Western blot. Jurkat cells (1E7) were treated for indicated time periods with 10 nm/ml PMA, 1 µM ionomycin and ab12134 (1/10000 dilution). 10 µl of cytoplasmic (C) and nuclear extract (N) were resolved on a 8.75% SDS-PAGE and transferred to Immobilon membrane.

ab12134 detects both phosphorylated and non-phosphorylated forms of IKB alpha by Western blot. Jurkat cells (1E7) were treated for indicated time periods with 10 nm/ml PMA, 1 µM ionomycin and ab12134 (1/10000 dilution). 10 µl of cytoplasmic (C) and nuclear extract (N) were resolved on a 8.75% SDS-PAGE and transferred to Immobilon membrane.