Phospho-STAT3 (S727) Recombinant Rabbit Monoclonal Antibody [SY24-09]
cat.: ET1607-39
Product Type: Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity: Human, Rat, Mouse
Applications: WB, IF-Cell, IF-Tissue, IHC-P, IP
Clonality: Monoclonal
Clone number: SY24-09
Form: Liquid
Storage condition: Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.
Storage buffer: 1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Concentration: 1ug/ul
Purification: Protein A affinity purified.
Molecular weight: Predicted band size: 88 kDa
Isotype: IgG
Immunogen: Synthetic phospho-peptide corresponding to residues surrounding Ser727 of Human STAT3 aa 701-750 / 770.
Positive control: Hela cell lysate, NIH/3T3 cell lysate, C6 cell lysate, C6 treated with 200ng/mL TPA for 35 minutes cell lysate, Rat cerebellum tissue lysate, Hela, NIH/3T3, Huh7, rat hippocampus tissue, rat brain tissue, human kidney tissue, mouse brain tissue, mouse liver tissue.
Subcellular location: Cytoplasm, Nucleus.
Recommended Dilutions:
  WB
  IF-Cell
  IF-Tissue
  IHC-P
  IP
1:1,000-1:2,000
1:100-1:200
1:200-1:500
1:500-1:1,000
Use at an assay dependent concentration.
Uniprot #: SwissProt: P40763 Human | P42227 Mouse | P52631 Rat
Alternative names: 1110034C02Rik Acute Phase Response Factor Acute-phase response factor ADMIO APRF AW109958 DNA binding protein APRF FLJ20882 HIES MGC16063 Signal transducer and activator of transcription 3 (acute phase response factor) Signal transducer and activator of transcription 3 STAT 3 Stat3 STAT3_HUMAN
Images
ET1607-39_1.jpg Fig1: Western blot analysis of Phospho-STAT3(S727) on Hela cell lysate with Rabbit anti-Phospho-STAT3(S727) antibody (ET1607-39) at 1/1,000 dilution.


Lysates/proteins at 15 µg/Lane.
Exposure time: 30 seconds; ECL: K1801


Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary antibody: ET1607-39, 1/1,000 in 5% NFDM/TBST, overnight at 4 ℃
Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature

Predicted band size: 88.1 kDa
Observed band size: 88 kDa
ET1607-39_2.jpg Fig2: Western blot analysis of Phospho-STAT3(S727) on Hela cell lysate with Rabbit anti-Phospho-STAT3(S727) antibody (ET1607-39) at 1/2,000 dilution.

Lane 1: NIH/3T3 cell lysate
Lane 2: C6 cell lysate

Lysates/proteins at 20 µg/Lane.
Exposure time: 30 seconds; ECL: K1801


Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary antibody: ET1607-39, 1/2,000 in 5% NFDM/TBST, overnight at 4 ℃
Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature

Predicted band size: 88.1 kDa
Observed band size: 88 kDa
ET1607-39_3.jpg Fig3: Western blot analysis of Phospho-STAT3 (S727) on different lysates with Rabbit anti-Phospho-STAT3 (S727) antibody (ET1607-39) at 1/1,000 dilution.

Lane 1: PC-12 cell lysate (20 µg/Lane)
Lane 2: C6 cell lysate (20 µg/Lane)
Lane 3: C6 treated with 200ng/mL TPA for 35 minutes cell lysate (20 µg/Lane)
Lane 4: Rat cerebellum tissue lysate (40 µg/Lane)

Predicted band size: 88 kDa
Observed band size: 88 kDa

Exposure time: 1 minute 50 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 (ET1607-39) 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.
ET1607-39_4.jpg Fig4: ICC staining of Phospho-STAT3 (S727) 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 (ET1607-39, 1/100) 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).
ET1607-39_5.jpg Fig5: ICC staining of Phospho-STAT3 (S727) 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 (ET1607-39, 1/100) 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).
ET1607-39_6.jpg Fig6: Immunocytochemistry analysis of Huh7 cells labeling Phospho-STAT3 (S727) with Rabbit anti-Phospho-STAT3 (S727) antibody (ET1607-39) 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-STAT3 (S727) antibody (ET1607-39) 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.
ET1607-39_7.jpg Fig7: Immunohistochemical analysis of paraffin-embedded rat hippocampus tissue with Rabbit anti-Phospho-STAT3 (S727) antibody (ET1607-39) at 1:500 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 (ET1607-39) at 1:500 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.
ET1607-39_8.jpg Fig8: Immunohistochemical analysis of paraffin-embedded rat brain tissue with Rabbit anti-Phospho-STAT3 (S727) antibody (ET1607-39) at 1:500 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 (ET1607-39) at 1:500 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.
ET1607-39_9.jpg Fig9: Immunohistochemical analysis of paraffin-embedded human kidney tissue with Rabbit anti-Phospho-STAT3 (S727) antibody (ET1607-39) at 1:500 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 (ET1607-39) at 1:500 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.
ET1607-39_10.jpg Fig10: Immunohistochemical analysis of paraffin-embedded mouse brain tissue with Rabbit anti-Phospho-STAT3 (S727) antibody (ET1607-39) 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 (ET1607-39) 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.
ET1607-39_11.jpg Fig11: Immunohistochemical analysis of paraffin-embedded mouse liver tissue with Rabbit anti-Phospho-STAT3 (S727) antibody (ET1607-39) 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 (ET1607-39) 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.
Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.