All lanes : Anti-CBL antibody [YE323] - C-terminal (ab32027) at 1/1000 dilution

Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : CBL knockout HEK293T cell lysate
Lane 3 : THP-1 cell lysate
Lane 4 : Raji 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: 100 kDa
Observed band size: 110 kDa why is the actual band size different from the predicted?

Lanes 1-4: Merged signal (red and green). Green - ab32027 observed at 110 kDa. Red - loading control ab8245 observed at 36 kDa.

 ab32027 Anti-CBL antibody [YE323] - C-terminal was shown to specifically react with CBL in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab267245 (knockout cell lysate ab257200) was used. Wild-type and CBL knockout samples were subjected to SDS-PAGE. ab32027 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: 10 bp deletion in exon9

Representative images of CBL knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using an EVOS M5000 microscope.