Human DDIT3 knockout HeLa cell lysate

• WB

Supplier Data

Western blot - Human DDIT3 knockout HeLa cell lysate (AB256889)

All lanes : Anti-DDIT3 antibody [9C8] (ab11419)

Lane 1 : Wild-type HeLa Vehicle Control Tunicamycin (20ug/mL, 4h) cell lysate
Lane 2 : Wild-type HeLa Treated Tunicamycin (20ug/mL, 4h) cell lysate
Lane 3 : DDIT3 knockout HeLa Vehicle Control Tunicamycin (20ug/mL, 4h) cell lysate
Lane 4 : DDIT3 knockout HeLa Treated Tunicamycin (20ug/mL, 4h) cell lysate

Performed under reducing conditions.

Observed band size : 25 kDa

False colour image of Western blot : Anti-DDIT3 antibody [9C8] staining at 5 μg/ml, shown in green; Rabbit anti-alpha Tubulin antibody [EP1332Y] (ab52866) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab11419 was shown to bind specifically to DDIT3. A band was observed at 25 kDa in wild-type y cell lysates with no signal observed at this size in DDIT3 knockout cell line ab265760 (knockout cell lysate ab256889). To generate this image, wild-type and DDIT3 knockout y cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution 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-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) at 1/20000 dilution.

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• WB

Lab

Western blot - Human DDIT3 knockout HeLa cell lysate (AB256889)

False colour image of Western blot : Anti-DDIT3 antibody [9C8] staining at 5 µg/ml, shown in green; Rabbit anti-alpha Tubulin antibody [EP1332Y] (ab52866) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab11419 was shown to bind specifically to DDIT3. A band was observed at 25 kDa in wild-type y cell lysates with no signal observed at this size in DDIT3 knockout cell line ab265760 (knockout cell lysate ab256889). To generate this image, wild-type and DDIT3 knockout y cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution 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-Mouse IgG H&L (IRDye^® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preabsorbed (ab216777) at 1/20000 dilution.

All lanes:

Western blot - Anti-CHOP antibody [9C8] (<a href='/en-us/products/primary-antibodies/chop-antibody-9c8-ab11419'>ab11419</a>) at 5 µg/mL

Lane 1:

Wild-type HeLa Vehicle Control Tunicamycin cell lysate at 20 µg

Lane 2:

Wild-type HeLa Treated Tunicamycin cell lysate at 20 µg

Lane 3:

DDIT3 knockout HeLa Vehicle Control Tunicamycin cell lysate at 20 µg

Lanes 3 - 4:

Western blot - Human DDIT3 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-ddit3-knockout-hela-cell-line-ab265760'>ab265760</a>)

Lanes 3 - 4:

Western blot - Human DDIT3 knockout HeLa cell lysate (ab256889)

Lane 4:

DDIT3 knockout HeLa Treated Tunicamycin cell lysate at 20 µg

Predicted band size: 19 kDa

Observed band size: 25 kDa

false

• WB

Supplier Data

Western blot - Human DDIT3 knockout HeLa cell lysate (AB256889)

Lanes 1 - 5 : Merged signal (red and green). Green - ab11419 observed at 26 kDa. Red - loading control ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) observed at 55kDa.

ab11419 was shown to react with DDIT3 in wild-type SW480 cells in western blot with loss of signal observed in DDIT3 knockout cell line ab269585 (knockout cell lysate ab270708). Wild-type and DDIT3 knockout SW480 cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab11419 and ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) overnight at 4°C at 5 µg/ml and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye^® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-CHOP antibody [9C8] (<a href='/en-us/products/primary-antibodies/chop-antibody-9c8-ab11419'>ab11419</a>) at 5 µg/mL

Lane 1:

Wild-type HeLa Vehicle Control Tunicamycin (20ug/mL, 4h) cell lysate at 20 µg

Lane 2:

Wild-type HeLa Treated Tunicamycin (20ug/mL, 4h) cell lysate at 20 µg

Lane 3:

DDIT3 knockout HeLa Vehicle Control Tunicamycin (20ug/mL, 4h) cell lysate at 20 µg

Lanes 3 - 4:

Western blot - Human DDIT3 knockout SW480 cell line (<a href='/en-us/products/cell-lines/human-ddit3-knockout-sw480-cell-line-ab269585'>ab269585</a>)

Lanes 3 - 4:

Western blot - Human DDIT3 knockout HeLa cell lysate (ab256889)

Lane 4:

DDIT3 knockout HeLa Treated Tunicamycin (20ug/mL, 4h) cell lysate at 20 µg

Predicted band size: 19 kDa

Observed band size: 25 kDa

false

• Sanger seq

Unknown

Sanger Sequencing - Human DDIT3 knockout HeLa cell lysate (AB256889)

Homozygous : 1 bp insertion in exon2