ERK1/2 Recombinant Rabbit Monoclonal Antibody [SA43-03] - BSA and Azide free

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Western blot analysis of ERK1/2 on different lysates with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/5,000 dilution and competitor's antibody at 1/1,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: Jurkat cell lysate
Lane 3: HepG2 cell lysate
Lane 4: NIH/3T3 cell lysate
Lane 5: PC-12 cell lysate
Lane 6: C6 cell lysate

Lysates/proteins at 15 µg/Lane.

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

Exposure time: 19 seconds; ECL: K1801;

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 (ET1601-29) at 1/5,000 dilution and competitor's antibody at 1/1,000 dilution were 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.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Immunocytochemistry analysis of Jurkat cells labeling ERK1/2 with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/2,000 dilution and competitor's antibody at 1/1,600 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/2,000 dilution and competitor's antibody at 1/1,600 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

☑ Knockdown (KD)

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Western blot analysis of ERK1/2 on different lysates with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/5,000 dilution.

Lane 1: HEK-293-si NT cell lysate
Lane 2: HEK-293-si ERK1/2 cell lysate

Lysates/proteins at 10 µg/Lane.

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

Exposure time: 20 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary antibody (ET1601-29, 1/5,000) and Loading control antibody (Rabbit anti-GAPDH, ET1601-4, 1/10,000) were 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.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Flow cytometric analysis of Jurkat cells labeling ERK1/2.

Cells were fixed and permeabilized. Then stained with the primary antibody (ET1601-29, red) at 1/1,000 dilution and competitor's antibody (red) at 1/800 dilution, compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded human breast carcinoma tissue with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/20,000 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 (ET1601-29) at 1/20,000 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.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded mouse esophagus tissue with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/20,000 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 (ET1601-29) at 1/20,000 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.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded mouse stomach tissue with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/20,000 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 (ET1601-29) at 1/20,000 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.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded rat esophagus tissue with Rabbit anti-ERK1/2 antibody (ET1601-29) 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 (ET1601-29) 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.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Immunohistochemical analysis of paraffin-embedded rat stomach tissue with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/3,000 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 (ET1601-29) at 1/3,000 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.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Application: IHC-Fr

Species: Mouse

Site: Hippocampus

Sample: Frozen section

Antibody concentration: 1/200

Antigen retrieval: The section was pre-treated using 1% SDS buffer (in PBS, pH 7.4) for 5 minutes at room temperature.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Application: IHC-Fr

Species: Mouse

Site: Cerebral cortex

Sample: Frozen section

Antibody concentration: 1/200

Antigen retrieval: The section was pre-treated using 1% SDS buffer (in PBS, pH 7.4) for 5 minutes at room temperature.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
ERK1/2 was immunoprecipitated in 0.2mg Jurkat cell lysate with ET1601-29 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using ET1601-29 at 1/5,000 dilution. Anti-Rabbit IgG for IP, AlpSdAbs® VHH(HRP) (025-100-005) at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: Jurkat cell lysate (input)
Lane 2: Rabbit IgG instead of ET1601-29 in Jurkat cell lysate
Lane 3: ET1601-29 IP in Jurkat cell lysate

Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 43 seconds; ECL: K1801

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Immunocytochemistry analysis of PC-12 cells labeling ERK1/2 with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Immunocytochemistry analysis of NIH/3T3 cells labeling ERK1/2 with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/500 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-ERK1/2 antibody (ET1601-29) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

This data was developed using ET1601-29, the same antibody clone in a different buffer formulation.
Immunoprecipitation (IP)

ERK1/2 was immunoprecipitated in 0.2 mg RAW264.7 (Mouse monocytic macrophage leukemia cell) cell lysate with ET1601-29 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using ET1601-29 at 1/5,000 dilution. Anti-Rabbit IgG for IP, AlpSdAbs® VHH(HRP) (025-100-005) at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: RAW264.7 cell lysate (input)
Lane 2: ET1601-29 IP in RAW264.7 cell lysate
Lane 3: Rabbit IgG instead of ET1601-29 in RAW264.7 cell lysate

Exposure time: 7 seconds
Blocking: 5% NFDM/TBST, 1 hour at room temperature

Primary dilution: ET1601-29, 1/5,000 in primary antibody dilution buffer (K1803), 2 hours at room temperature

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