Anti-ATF-4 antibody [EPR30044-580] - BSA and Azide free

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-ATF-4 antibody [EPR30044-580] - BSA and Azide free (AB325277)

This data was developed using ab325275, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized HeLa (human cervical adenocarcinoma epithelial cell); HeLa treated with tunicamycin (2 ug/mL) for 8 hr cells labelling ATF-4 with ab325275 at 1/50 (10.24 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/ab150081 1000 2ug/ml dilution (Green).

Confocal image showing increased nuclear staining in HeLa cells treated with tunicamycin (2 ug/mL) for 8 hr (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/ab7291 1000 1ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/ab150081 1000 2ug/ml dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-ATF-4 antibody [EPR30044-580] - BSA and Azide free (AB325277)

This data was developed using ab325275, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage); RAW 264.7 treated with tunicamycin (2 ug/mL) for 8 hr cells labelling ATF-4 with ab325275 at 1/50 (10.24 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/ab150081 1000 2ug/ml dilution (Green).

Confocal image showing increased nuclear staining in RAW 264.7 cells treated with tunicamycin (2 ug/mL) for 8 hr (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/ab7291 1000 1ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/ab150081 1000 2ug/ml dilution.

• WB

Supplier Data

Western blot - Anti-ATF-4 antibody [EPR30044-580] - BSA and Azide free (AB325277)

This data was developed using ab325275, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression of ATF-4 is upregulated in response to Tunicamycin treatment (PMID : 30926605).

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-ATF-4 antibody [EPR30044-580] (<a href='/en-us/products/primary-antibodies/atf-4-antibody-epr30044-580-ab325275'>ab325275</a>) at 1/1000 dilution

Lane 1:

Untreated HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 50 µg

Lane 2:

HeLa treated with 2 μg/ml Tunicamycin for 8 hours whole cell lysate at 50 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 50 kDa,124 kDa

false

Exposure time: 48s

• WB

Supplier Data

Western blot - Anti-ATF-4 antibody [EPR30044-580] - BSA and Azide free (AB325277)

This data was developed using ab325275, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

The expression of ATF-4 is upregulated in response to Tunicamycin treatment (PMID : 30926605).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-ATF-4 antibody [EPR30044-580] (<a href='/en-us/products/primary-antibodies/atf-4-antibody-epr30044-580-ab325275'>ab325275</a>) at 1/1000 dilution

Lane 1:

Untreated RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Lane 2:

RAW 264.7 treated with 2 μg/ml Tunicamycin for 8 hours whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 50 kDa,36 kDa

false

Exposure time: 81s

• WB

Supplier Data

Western blot - Anti-ATF-4 antibody [EPR30044-580] - BSA and Azide free (AB325277)

This data was developed using ab325275, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

This antibody does not cross-react with mouse ATF-5.

In Western blot, Anti-ATF-4/5 antibody staining at 1/1000 dilution.

All lanes:

Western blot - Anti-ATF-4 antibody [EPR30044-580] (<a href='/en-us/products/primary-antibodies/atf-4-antibody-epr30044-580-ab325275'>ab325275</a>) at 1/1000 dilution

Lane 1:

Untagged mouse AFT-4 recombinant protein at 10 ng

Lane 2:

His-tagged mouse AFT-5 recombinant protein at 100 ng

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 50 kDa,32 kDa

false

Exposure time: 15s