All lanes : Anti-A RAF antibody [EPR16208] (ab200653) at 1/1000 dilution

Lane 1 : Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
Lane 2 : ARAF knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
Lane 3 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 4 : Raji (Human Burkitts lymphoma cell line) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution

Predicted band size: 68 kDa
Observed band size: 68 kDa

Lanes 1-4: Merged signal (red and green). Green - ab200653 observed at 68 kDa. Red - loading control ab8245 observed at 36 kDa.

 ab200653 Anti-A RAF antibody [EPR16208] was shown to specifically react with A RAF in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266351 (knockout cell lysate ab257838) was used. Wild-type and A RAF knockout samples were subjected to SDS-PAGE. ab200653 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.

Homozygous: 1 bp deletion in exon 2