All lanes : Anti-ATG16L1 antibody [5H9A11] (ab233796) at 1/1000 dilution

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

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 68 kDa
Observed band size: 68 kDa

Lanes 1- 4: Merged signal (red and green). Green - ab233796 observed at 68 kDa. Red - Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) observed at 37 kDa.

 ab233796 was shown to 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 HeLa and ATG16L1 knockout HeLa cell lysates 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) overnight at 4°C at a 1 in 1000 dilution and a 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.

All lanes : Anti-ATG16L1 antibody [5H9A11] (ab233796) at 1/500 dilution

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

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 68 kDa

Lanes 1-4: Merged signal (red and green). Green - ab233796 observed at 68 and 72 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. Loss of signal was observed when knockout cell line ab261773 (knockout cell lysate ab256844) was used. Wild-type and ATG16L1 knockout samples were subjected to SDS-PAGE. ab233796 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-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.

 

 

All lanes : Anti-ATG16L1 antibody [5H9A11] (ab233796) at 1/500 dilution

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

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 68 kDa

Lanes 1-4: Merged signal (red and green). Green - ab233796 observed at 68 and 72 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. Loss of signal was observed when knockout cell line ab261772 (knockout cell lysate ab256843) was used. Wild-type and ATG16L1 knockout samples were subjected to SDS-PAGE. ab233796 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-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.

Anti-ATG16L1 antibody [5H9A11] (ab233796) at 1/500 dilution + Recombinant human ATG16L1 (aa 11-257) protein

Predicted band size: 68 kDa

Expected MW is 56 kDa.

All lanes : Anti-ATG16L1 antibody [5H9A11] (ab233796) at 1/500 dilution

Lane 1 : Untransfected HEK-293 (human epithelial cell line from embryonic kidney) cell lysate
Lane 2 : ATG16L1 (aa 11-257)-hIgG-Fc-transfected HEK-293 cell lysate

Predicted band size: 68 kDa

All lanes : Anti-ATG16L1 antibody [5H9A11] (ab233796) at 1/500 dilution

Lane 1 : HeLa (human epithelial cell line from cervix adenocarcinoma) cell lysate
Lane 2 : Raji (human Burkitt's lymphoma cell line) cell lysate
Lane 3 : PANC-1 (human pancreatic epithelial cancinoma cell line) cell lysate
Lane 4 : Jurkat (human T cell leukemia cell line from peripheral blood) cell lysate
Lane 5 : PC-12 (rat adrenal gland pheochromocytoma cell line) cell lysate
Lane 6 : HepG2 (human liver hepatocellular carcinoma cell line) cell lysate
Lane 7 : HEK-293 (human epithelial cell line from embryonic kidney) cell lysate
Lane 8 : NIH/3T3 (mouse embryo fibroblast cell line) cell lysate

Predicted band size: 68 kDa

Flow cytometric analysis of HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling ATG16L1 with ab233796 at 1/200 dilution (green) compared with a negative control (red).