All lanes : Anti-FZR1 antibody [AR38.2] (ab77885) at 1/500 dilution

Lane 1 : Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
Lane 2 : FZR1 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 : A431 (Human epidermoid carcinoma cell line) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) at 1/10000 dilution

Predicted band size: 55 kDa
Observed band size: 55 kDa

Lanes 1-4: Merged signal (red and green). Green - ab77885 observed at 55 kDa. Red - loading control ab181602 observed at 36 kDa.

 ab77885 Anti-FZR1 antibody [AR38.2] was shown to specifically react with FZR1 in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab267263 (knockout cell lysate ab263208) was used. Wild-type and FZR1 knockout samples were subjected to SDS-PAGE. ab77885 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) were incubated overnight at 4°C at 1 in 500 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye^® 800CW) preadsorbed (ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

Homozygous: 1 bp insertion in exon9

Representative images FZR1 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 a EVOS M5000 microscope.