All lanes : Anti-Met (c-Met) antibody [EP1454Y] - N-terminal (ab51067) at 1/1000 dilution

Lane 1 : Wild-type HeLa
Lane 2 : MET knockout HeLa

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/20000 dilution

Predicted band size: 155 kDa
Observed band size: 37 kDa why is the actual band size different from the predicted?

Lanes 1 - 2: Merged signal (red and green). Green - ab51067 observed at 156 kDa. Red - loading control, ab8245 observed at 37 kDa.  

 ab51067 was shown to react with Met (c-Met) in wild-type HeLa. Loss of signal was observed when knockout cell line ab265961 (knockout cell lysate ab256991) was used. Wild-type and Met (c-Met) knockout samples were subjected to SDS-PAGE. ab51067 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: 5 bp deletion in exon 2.

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