Phospho-FAK (Y397) Recombinant Rabbit Monoclonal Antibody [SC54-07]

Western blot analysis of Phospho-FAK (Y397) on different lysates with Rabbit anti-Phospho-FAK (Y397) antibody (ET1610-34) at 1/500 dilution.

Lane 1: Hela cell lysate
Lane 2: PC-3 cell lysate

Lysates/proteins at 10 µg/Lane.

Predicted band size: 119 kDa
Observed band size: 119 kDa

Exposure time: 2 minutes;

6% 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 (ET1610-34) at 1/500 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:300,000 dilution was used for 1 hour at room temperature.

☑ Cell treatment (CT)

Western blot analysis of Phospho-FAK (Y397) on Hela cell lysates.

Lane 1: Hela cells, whole cell lysate, 10ug/lane
Lane 2: Hela cells treated with 2.8ug/ul lambda-PP for 30 minutes, whole cell lysates, 10ug/lane

All lanes :
Anti-Phospho-FAK (Y397) antibody (ET1610-34) at 1:500 dilution. Anti-Hsp90 antibody (ET1605-56) at 1:10,000 dilution. Goat Anti-Rabbit IgG H&L (HRP) (HA1001) at 1/200,000 dilution.

Predicted band size: 119 kDa
Observed band size: 119 kDa

Blocking and diluting buffer: 5% BSA.

Exposure time: 30 seconds

Immunocytochemistry analysis of NIH/3T3 cells labeling Phospho-FAK (Y397) with Rabbit anti-Phospho-FAK (Y397) antibody (ET1610-34) at 1/50 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 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-Phospho-FAK (Y397) antibody (ET1610-34) at 1/50 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 (HA601187, 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.

Immunocytochemistry analysis of C6 cells labeling Phospho-FAK (Y397) with Rabbit anti-Phospho-FAK (Y397) antibody (ET1610-34) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 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-Phospho-FAK (Y397) antibody (ET1610-34) 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 (HA601187, 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.

ICC staining of Phospho-FAK (Y397) in Hela 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 (ET1610-34, 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).

Immunohistochemical analysis of paraffin-embedded mouse spleen tissue with Rabbit anti-Phospho-FAK (Y397) antibody (ET1610-34) at 1/200 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 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 (ET1610-34) 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 rat spleen tissue with Rabbit anti-Phospho-FAK (Y397) antibody (ET1610-34) at 1/200 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 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 (ET1610-34) 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.