Phospho-Erk1 (T202)+Erk2 (T185) Recombinant Rabbit Monoclonal Antibody [SZ2-4]

☑ Cell treatment (CT)

Western blot analysis of Phospho-Erk1 (T202)+Erk2 (T185) on different lysates with Rabbit anti-Phospho-Erk1 (T202)+Erk2 (T185) antibody (ET1603-22) at 1/1,000 dilution.

Lane 1: Jurkat cell lysate
Lane 2: Jurkat treated with 200ng/mL PMA for 35 minutes cell lysate
Lane 3: NIH/3T3 cell lysate
Lane 4: NIH/3T3 treated with 200nM PMA for 30 minutes cell lysate
Lane 5: C6 cell lysate
Lane 6: C6 treated with 200nM PMA for 30 minutes cell lysate

Lysates/proteins at 10 µg/Lane.

Predicted band size: 41/43 kDa
Observed band size: 41/43 kDa
Exposure time: 3 minutes; ECL: K1802;
4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1603-22) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

☑ Cell treatment (CT)

Western blot analysis of Phospho-Erk1(T202)+Erk2(T185) on jurkat cell lysates.

Lane 1: jurkat cells, whole cell lysate, 10ug/lane
Lane 2/3: jurkat cells treated with 200 ng/ml PMA for 35 minutes, whole cell lysate, 10ug/lane
Lane 4: jurkat cells treated with 200 ng/ml PMA for 35 minutes, then treated with 2.8ug/ul lambda-PP for 30 minutes, whole cell lysates, 10ug/lane

All lanes :
Anti-Phospho-Erk1(T202)+Erk2(T185) antibody (ET1603-22) at 1/500 dilution. Anti-Erk1+Erk2antibody (ET1601-29) at 1/500 dilution. Anti-GAPDH antibody (ET1601-4) at 1/10,000 dilution. Goat Anti-Rabbit IgG H&L (HRP) (HA1001) at 1/200,000 dilution.

Predicted band size: 41/43 kDa
Observed band size: 41/43 kDa

Blocking and diluting buffer: 5% BSA.
Exposure time: Lan1/2 4 minutes; Lan3/4 3 minutes

ICC staining of Phospho-Erk1 (T202)+Erk2 (T185) in A549 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1603-22, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).

ICC staining of Phospho-Erk1 (T202)+Erk2 (T185) in NIH/3T3 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1603-22, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).

ICC staining of Phospho-Erk1 (T202)+Erk2 (T185) in MCF-7 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1603-22, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).

Flow cytometric analysis of Phospho-Erk1 (T202)+Erk2 (T185) was done on MCF-7 cells. The cells were fixed, permeabilized and stained with the primary antibody (ET1603-22, 1/50) (red). After incubation of the primary antibody at room temperature for an hour, the cells were stained with a Alexa Fluor 488-conjugated Goat anti-Rabbit IgG Secondary antibody at 1/1000 dilution for 30 minutes.Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

Immunohistochemical analysis of paraffin-embedded human gallbladder tissue with Rabbit anti-Phospho-Erk1 (T202)+Erk2 (T185) antibody (ET1603-22) at 1/100 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1603-22) at 1/100 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

Immunohistochemical analysis of paraffin-embedded human kidney tissue with Rabbit anti-Phospho-Erk1 (T202)+Erk2 (T185) antibody (ET1603-22) at 1/200 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1603-22) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX."

Immunohistochemical analysis of paraffin-embedded human spleen tissue with Rabbit anti-Phospho-Erk1 (T202)+Erk2 (T185) antibody (ET1603-22) at 1/200 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1603-22) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX."

Immunohistochemical analysis of paraffin-embedded mouse kidney tissue with Rabbit anti-Phospho-Erk1 (T202)+Erk2 (T185) antibody (ET1603-22) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1603-22) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX."

Immunohistochemical analysis of paraffin-embedded mouse spleen tissue with Rabbit anti-Phospho-Erk1 (T202)+Erk2 (T185) antibody (ET1603-22) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1603-22) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX."

Immunohistochemical analysis of paraffin-embedded rat kidney tissue with Rabbit anti-Phospho-Erk1 (T202)+Erk2 (T185) antibody (ET1603-22) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1603-22) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX."

Immunohistochemical analysis of paraffin-embedded rat spleen tissue with Rabbit anti-Phospho-Erk1 (T202)+Erk2 (T185) antibody (ET1603-22) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1603-22) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX."

Immunohistochemical analysis of paraffin-embedded human thyroid cancer tissue with Rabbit anti-Phospho-Erk1 (T202)+Erk2 (T185) antibody (ET1603-22) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1603-22) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX."