Anti-ATG16L1 antibody [EPR15638] - BSA and Azide free

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATG16L1 antibody [EPR15638] - BSA and Azide free (AB232636)

Immunohistochemical analysis of paraffin-embedded Mouse colon tissue labeling ATG16L1 with ab187671 at 1/100 dilution followed by pre-diluted HRP Polymer for Rabbit IgG secondary antibody and counter-stained with Hematoxylin.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab187671).

Perform heat mediated antigen retrieval with EDTA buffer pH 9 before commencing with IHC staining protocol.

• WB

Lab

Western blot - Anti-ATG16L1 antibody [EPR15638] - BSA and Azide free (AB232636)

False colour image of Western blot : Anti-ATG16L1 antibody [EPR15638] - N-terminal staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab187671 was shown to bind specifically to ATG16L1. A band was observed at 68, 70 kDa in wild-type THP-1 cell lysates with no signal observed at this size in ATG16L1 knockout cell line ab277834 (knockout cell lysate ab278184). To generate this image, wild-type and ATG16L1 knockout THP-1 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-ATG16L1 antibody [EPR15638] - N-terminal (<a href='/en-us/products/primary-antibodies/atg16l1-antibody-epr15638-n-terminal-ab187671'>ab187671</a>) at 1/1000 dilution

Lane 1:

Wild-type THP-1 cell lysate at 20 µg

Lane 2:

Western blot - Human ATG16L1 knockout THP-1 cell lysate (<a href='/en-us/products/cell-lysates/human-atg16l1-knockout-thp-1-cell-lysate-ab278184'>ab278184</a>) at 20 µg

Lane 3:

Wild type HeLa cell lysate at 20 µg

Lane 4:

ATG16L1 knockout HeLa cell lysate at 20 µg

Predicted band size: 68 kDa

Observed band size: 68 kDa,70 kDa

false

• WB

Supplier Data

Western blot - Anti-ATG16L1 antibody [EPR15638] - BSA and Azide free (AB232636)

This data was developed using the same antibody clone in a different buffer formulation (ab187671).

Lanes 1-4 : Merged signal (red and green). Green - ab187671 observed at 68 and 72 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. 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. ab187671 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.

All lanes:

Western blot - Anti-ATG16L1 antibody [EPR15638] - N-terminal (<a href='/en-us/products/primary-antibodies/atg16l1-antibody-epr15638-n-terminal-ab187671'>ab187671</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

ATG16L1 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human ATG16L1 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-atg16l1-knockout-hela-cell-line-ab261772'>ab261772</a>)

Lane 3:

Jurkat cell lysate at 20 µg

Lane 4:

Daudi cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Predicted band size: 68 kDa

Observed band size: 68 kDa,72 kDa

false

• WB

Lab

Western blot - Anti-ATG16L1 antibody [EPR15638] - BSA and Azide free (AB232636)

This data was developed using the same antibody clone in a different buffer formulation (ab187671).

Lanes 1-4 : Merged signal (red and green). Green - ab187671 observed at 75 kDa. Red - loading control ab8245 observed at 36 kDa.

ab187671 Anti-ATG16L1 antibody [EPR15638] 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.

All lanes:

Western blot - Anti-ATG16L1 antibody [EPR15638] - N-terminal (<a href='/en-us/products/primary-antibodies/atg16l1-antibody-epr15638-n-terminal-ab187671'>ab187671</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

ATG16L1 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human ATG16L1 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-atg16l1-knockout-hela-cell-line-ab265263'>ab265263</a>)

Lane 3:

Jurkat cell lysate at 20 µg

Lane 4:

Daudi cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 68 kDa

Observed band size: 75 kDa

false

• WB

Lab

Western blot - Anti-ATG16L1 antibody [EPR15638] - BSA and Azide free (AB232636)

This data was developed using the same antibody clone in a different buffer formulation (ab187671).

Lanes 1-4 : Merged signal (red and green). Green - ab187671 observed at 68 and 72 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. 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. ab187671 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.

All lanes:

Western blot - Anti-ATG16L1 antibody [EPR15638] - N-terminal (<a href='/en-us/products/primary-antibodies/atg16l1-antibody-epr15638-n-terminal-ab187671'>ab187671</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

ATG16L1 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human ATG16L1 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-atg16l1-knockout-hela-cell-line-ab261773'>ab261773</a>)

Lane 3:

Jurkat cell lysate at 20 µg

Lane 4:

Daudi cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Predicted band size: 68 kDa

Observed band size: 68 kDa,72 kDa

false

• WB

Supplier Data

Western blot - Anti-ATG16L1 antibody [EPR15638] - BSA and Azide free (AB232636)

Lanes 1 - 4 : Merged signal (red and green). Green - ab187671 observed at 68 and 70 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab187671 was shown to recognize ATG16L1 when ATG16L1 knockout samples were used, along with additional cross-reactive bands. Wild-type and ATG16L1 knockout samples were subjected to SDS-PAGE. ab187671 and ab8245 (loading control to GAPDH) were diluted 1/2000 and 1/10000 respectively and incubated overnight at 4°C. 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 1/10000 dilution for 1 h at room temperature before imaging.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab187671).

All lanes:

Western blot - Anti-ATG16L1 antibody [EPR15638] - BSA and Azide free (ab232636) at 1/2000 dilution

Lane 1:

Wild-type HAP1 cell lysate at 20 µg

Lane 2:

ATG16L1 knockout HAP1 cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

Jurkat cell lysate at 20 µg

Predicted band size: 68 kDa

false