Anti-p53 (phospho S392) antibody [EP155Y]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human lung carcinoma tissue sections labeling p53 with Purified ab33889 at 1 : 250 dilution. Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used at 1 : 0 dilution. PBS instead of the primary antibody was used as the negative control.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

Unpurified ab33889 at a 1/100 dilution staining p53 in paraffin embedded human prostate adenocarcinoma tissue by Immunohistochemistry.

• IP

Lab

Immunoprecipitation - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

ab33889 (purified) at 1 : 20 dilution (0.6μg) immunoprecipitating p53 in 293T whole cell lysate.

Lane 1 (input) : 293T (Human embryonic kidney epithelial cell) whole cell lysate, 10μg
Lane 2 (+) : ab33889 & 293T whole cell lysate
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab33889 in 293T whole cell lysate.

For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1 : 1000 dilution.
Blocking and diluting buffer : 5% NFDM/TBST.

All lanes:

Immunoprecipitation - Anti-p53 (phospho S392) antibody [EP155Y] (ab33889)

Predicted band size: 43 kDa

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• WB

Lab

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

Blocking and dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (ab33889) at 1/2000 dilution

Lane 1:

A431 whole cell lysate at 10 µg

Lane 2:

A431 treated with 1μg/ml doxorubicin for 24 hours whole cell lysate at 10 µg

Lane 3:

A431 treated with 1μg/ml doxorubicin for 24 hours whole cell lysate, the membrane was incubated with alkaline phosphatase at 10 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 1/100000 dilution

Predicted band size: 43 kDa

Observed band size: 53 kDa

false

Exposure time: 15s

• WB

Lab

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

Blocking and diluting buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (ab33889) at 1/1000 dilution

Lane 1:

HEK-293 (Human embryonic kidney epithelial cell) whole cell lysates at 15 µg

Lane 2:

HEK-293 (Human embryonic kidney epithelial cell) whole cell lysates, then the membrane was incubated with alkaline phosphatase at 15 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

Predicted band size: 43 kDa

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• WB

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Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

Lanes 1 and 5 : Extract of Hek293T incubated with etoposide (7.5 μg).
Lanes 2 and 6 : Lambda phosphatase (400 times-diluted)-treated extract of Hek293T incubated with etoposide (7.5 μg).
Lanes 3 and 7 : Lambda phosphatase (100 times-diluted)-treated extract of Hek293T incubated with etoposide (7.5 μg).
Lanes 4 and 8 : Lambda phosphatase (25 times-diluted)-treated extract of Hek293T incubated with etoposide (7.5 μg).

SDS PAGE performed under reducing conditions (100mM DTT, sample heated at 50°C).

Primary :
Lanes 1-4 : Anti p53 (phosphoS392) antibody (ab33889, unpurified) at 1/2000 dilution.
Lanes 5-8 : Anti p53 antibody (ab1101) at 1/2500 dilution.

Secondary :
Lanes 1-4 : HRP-conjugated goat anti-rabbit IgG (H&L) at 1/10000.
Lanes 5-8 : HRP-conjugated goat anti-mouse IgG (H&L) at 1/10000.

Blocked in 5% milk in PBS for 3 hours at room temperature.
Incubated with the primary antibody in 5% BSA + 50mM Tris pH 7.5 + 0.05% Tween-20 overnight at 4°C.
Incubated with the secondary antibody in blocking buffer for 2 hours at room temperature.

All lanes:

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (ab33889)

Predicted band size: 43 kDa

Observed band size: 53 kDa

true

Exposure time: 1min

• WB

Lab

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

Blocking and dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (ab33889) at 1/1000 dilution

Lane 1:

HEK-293 whole cell lysate - untreated at 10 µg

Lane 2:

HEK-293 whole cell lysate - treated with Alkaline Phosphatase at 10 µg

Secondary

All lanes:

Peroxidase-conjugated goat anti-rabbit IgG (H+L) at 1/1000 dilution

Predicted band size: 43 kDa

Observed band size: 43 kDa

false

Exposure time: 15s

• WB

Unknown

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

All lanes:

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (ab33889) at 1/500 dilution

Lane 1:

MCF7 cell lysate untreated.

Lane 2:

MCF7 cell lysate treated with 5 ug/ml Actinomycin for 3hrs.

Lane 3:

MCF7 cell lysate treated with 5 ug/ml Actinomycin for 6hrs.

Lane 4:

MCF7 cell lysate treated with 5 ug/ml Actinomycin for 18hrs.

Predicted band size: 43 kDa

Observed band size: 53 kDa

false

• WB

Lab

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

Blocking and diluting buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (ab33889) at 1/1000 dilution

Lane 1:

RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates at 15 µg

Lane 2:

RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates, then the membrane was incubated with alkaline phosphatase at 15 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

Predicted band size: 43 kDa

false

• WB

Lab

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

Blocking and diluting buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-p53 (phospho S392) antibody [EP155Y] (ab33889) at 1/1000 dilution

Lane 1:

PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysates at 15 µg

Lane 2:

PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysates, then the membrane was incubated with alkaline phosphatase. at 15 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

Predicted band size: 43 kDa

Observed band size: 53 kDa

false

• Dot

Unknown

Dot Blot - Anti-p53 (phospho S392) antibody [EP155Y] (AB33889)

Dot blot analysis of p53 (pS392) peptide (Lane 1) and p53 non-phospho peptide (Lane 2) labelling p53 (pS392) with unpurified ab33889 at a dilution of 1/1000. ab97051 (Peroxidase conjugated goat anti-rabbit IgG (H+L)) was used as the secondary antibody at a dilution of 1/100000.

Blocking and dilution buffer : 5% NFDM/TBST.

Exposure time : 3 minutes.