Lane 1: Wild-type HeLa cell lysate (20µg)

Lane 2: ATG16L1 knockout HeLa cell lysate (20µg)
Lane 3: Jurkat cell lysate (20µg)
Lane 4: Daudi cell lysate (20µg)
Lanes 1- 4: Merged signal (red and green). Green - ab233796 observed at 68 kDa. Red - loading control, ab181602 observed at 37 kDa.
ab233796 Anti-ATG16L1 antibody [5H9A11] was shown to specifically react with ATG16L1 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab265263 (knockout cell lysate ab256842) was used. Wild-type and ATG16L1 knockout samples were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab233796 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) were incubated overnight at 4^°C at 1 in 1000 and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye^® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye^® 680RD) preadsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

Lane 1: Wild-type HeLa cell lysate (20µg)

Lane 2: ATG16L1 knockout HeLa cell lysate (20µg)
Lane 3: Jurkat cell lysate (20µg)
Lane 4: Daudi cell lysate (20µg)
Lanes 1- 4: Merged signal (red and green). Green - ab187671 observed at 68 kDa. Red - loading control, ab8245 observed at 37 kDa.
ab187671 Anti-ATG16L1 antibody [EPR15638] - N-terminal was shown to specifically react with ATG16L1 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab265263 (knockout cell lysate ab256842) was used. Wild-type and ATG16L1 knockout samples were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab187671 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4^°C at 1 in 1000 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.

Lane 1:Wild-type HeLa cell lysate (20 ug)
Lane 2:ATG16L1 knockout HeLa cell lysate (20 ug)
Lane 3:Jurkat cell lysate (20 ug)
Lane 4:Daudi cell lysate (20 ug)

ab187671 was shown to specifically react with ATG16L1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265263 (knockout cell lysate ab256842) was used. Wild-type and ATG16L1 knockout samples were subjected to SDS-PAGE. ab187671 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2.5 hours 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: 1 bp insertion in exon1

 

Allele-2: Insertion of the selection cassette in exon1