All lanes : Anti-ATG10 antibody [EPR4804] (ab124711) at 1/1000 dilution

Lane 1 : Wild-type HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : ATG10 knockout HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 3 : K562 (Human chronic myelogenous leukemia lymphoblast cell line) whole cell lysate
Lane 4 : HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) 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: 25 kDa
Observed band size: 28 kDa why is the actual band size different from the predicted?

Lanes 1-4: Merged signal (red and green). Green - ab124711 observed at 28 kDa. Red - loading control ab8245 observed at 36 kDa.

 ab124711 Anti-ATG10 antibody [EPR4804] was shown to specifically react with ATG10 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265682 (knockout cell lysate ab258786) was used. Wild-type and ATG10 knockout samples were subjected to SDS-PAGE. ab124711 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.

Allele-1: 92 bp insertion in exon 5.

 

Allele-2: 5 bp deletion in exon 5.

 

Representative images ATG10 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 M5000 microscope.