Revision 17

#9145

Store at -20C

CST Logo
Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Applications:
W, W-F, W-S, IP, IHC-Bond, IHC-P, IF-IC, FC-FP, ChIP, ChIP-seq

Reactivity:
H M R Mk

Sensitivity:
Endogenous

MW (kDa):
79, 86

Source/Isotype:
Rabbit IgG

UniProt ID:
#P40763

Entrez-Gene Id:
6774

Product Usage Information

For optimal ChIP and ChIP-seq results, use 5 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.
Application Dilution
Western Blotting 1:2000
Fluorescent Western 1:2000
Simple Western™ 1:10 - 1:50
Immunoprecipitation 1:100
IHC Leica Bond 1:100 - 1:400
Immunohistochemistry (Paraffin) 1:100 - 1:400
Immunofluorescence (Immunocytochemistry) 1:100 - 1:400
Flow Cytometry (Fixed/Permeabilized) 1:100 - 1:400
Chromatin IP 1:100
Chromatin IP-seq 1:100

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

For a carrier-free (BSA and azide free) version of this product see product #73533.

Specificity/Sensitivity

Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody detects endogenous levels of Stat3 only when phosphorylated at tyrosine 705. This antibody does not cross-react with phospho-EGFR or the corresponding phospho-tyrosines of other Stat proteins.

Species predicted to react based on 100% sequence homology

Hamster, Bovine, Pig, Horse

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Tyr705 of mouse Stat3.

Background

The Stat3 transcription factor is an important signaling molecule for many cytokines and growth factor receptors (1) and is required for murine fetal development (2). Research studies have shown that Stat3 is constitutively activated in a number of human tumors (3,4) and possesses oncogenic potential (5) and anti-apoptotic activities (3). Stat3 is activated by phosphorylation at Tyr705, which induces dimerization, nuclear translocation, and DNA binding (6,7). Transcriptional activation seems to be regulated by phosphorylation at Ser727 through the MAPK or mTOR pathways (8,9). Stat3 isoform expression appears to reflect biological function as the relative expression levels of Stat3α (86 kDa) and Stat3β (79 kDa) depend on cell type, ligand exposure, or cell maturation stage (10). It is notable that Stat3β lacks the serine phosphorylation site within the carboxy-terminal transcriptional activation domain (8).

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

W: Western Blotting W-F: Fluorescent Western W-S: Simple Western™ IP: Immunoprecipitation IHC-Bond: IHC Leica Bond IHC-P: Immunohistochemistry (Paraffin) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized) ChIP: Chromatin IP ChIP-seq: Chromatin IP-seq

Cross-Reactivity Key

H: Human M: Mouse R: Rat Mk: Monkey

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 17

Cell Signaling Technology Logo
Western blot analysis of extracts from IFN-alpha treated Jurkat cells and HeLa cells (left), as well as EGF treated A431 cells (right), using Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody. Note that the basal phospho-Stat3 in A431 is detected by the antibody.
Western Blotting Image 1: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Western blot analysis of extracts from HeLa cells, untreated or treated with IFN-alpha (#36000, 100 ng/mL, 5 min) or IFN-gamma (#80385, 100 ng/mL, 30 min); using Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody #9145 (upper), Stat3 (D3Z2G) Rabbit Monoclonal Antibody #12640 (middle), and beta-Actin (D6A8) Rabbit Monoclonal antibody #8457 (lower).
Western Blotting Image 2: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Western blot analysis of extracts from HeLa cells, untreated (-) or treated with INF-alpha (100 ng/ml, 5 min; +), using Stat3 (124H6) Mouse Monoclonal Antibody #9139 (Panel A) and Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal #9145 (Panel B). Anti-mouse IgG (H+L) (DyLight® 680 Conjugate) #5470 (red) and Anti-rabbit IgG (H+L) (DyLight® 800 4X PEG Conjugate) #5151 (green) were used as secondary antibodies.
Western Blotting Image 1: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 17

Cell Signaling Technology Logo
Simple Western™ analysis of lysates (0.1 mg/mL) from serum-starved HeLa cells treated with IFN-alpha (100 ng/mL, 5 min) using Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody #9145. The virtual lane view (left) shows a single target band (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
Western Blotting Image 1: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Immunoprecipitation of phospho-Stat3 (Tyr705) from U266 extracts treated with human IFN-alpha (50 ng/ml, 15 min). Lane 1 is 10% input, lane 2 is Rabbit (DA1E) Monoclonal Antibody IgG Isotype Control #3900, and lane 3 is Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody. Western blot analysis was performed using Phospho-Stat3 (Tyr705) (3E2) Mouse Monoclonal Antibody #9138. Anti-mouse IgG, HRP-linked Antibody #7076 was used as a secondary antibody.
Immunoprecipitation Image 1: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Immunohistochemical analysis of paraffin-embedded human prostate adenocarcinoma using Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody performed on the Leica® BOND™ Rx.
Immunohistochemistry Image 1: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 17

Cell Signaling Technology Logo
Immunohistochemical analysis of paraffin-embedded human neuroendocrine lung carcinoma using Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody performed on the Leica® BOND™ Rx.
Immunohistochemistry Image 2: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Immunohistochemical analysis of paraffin-embedded Apc (min/+) mouse intestine, using Phosho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody.
Immunohistochemistry Image 1: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Immunnohistochemical analysis of paraffin-embedded human lung carcinoma, showing nuclear localization, using Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody.
Immunohistochemistry Image 2: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 17

Cell Signaling Technology Logo
Immunohistochemical analysis of paraffin embedded human breast carcinoma, specifically endothelial cells, untreated (left) or lambda phosphatase treated (right), using Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody.
Immunohistochemistry Image 3: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Immunohistochemical analysis using Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody on SignalSlide® HeLa -/+ IFNa IHC Controls #55861 (paraffin-embedded HeLa cell pellets, untreated (left) or treated with Human Interferon-α1 (hIFN-α1 (right)).
Immunohistochemistry Image 4: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Confocal immunofluorescent analysis of HeLa cells, IFN-alpha treated (left) or untreated (right), labeled with Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody (green).
Immunofluorescence Image 1: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 17

Cell Signaling Technology Logo
Flow cytometric analysis of U266 cells, untreated (blue) or treated with IFN-alpha (50 ng/ml, 15 min; green) using Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody (solid lines) or concentration matched Rabbit (DA1E) Monoclonal Antibody IgG Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Flow Cytometry Image 1: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Chromatin immunoprecipitations were performed with cross-linked chromatin from Hep G2 cells starved overnight and treated with IL-6 (100 ng/ml) for 30 minutes and Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across IRF1, a known target gene of Phospho-Sata3 (see additional figure containing ChIP-qPCR data). For additional ChIP-seq tracks, please download the product datasheet.
Chromatin Immunoprecipitation Image 1: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Chromatin immunoprecipitations were performed with cross-linked chromatin from Hep G2 cells starved overnight and treated with IL-6 (100 ng/ml) for 30 minutes and Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795. The figure shows binding across chromosome 5 (upper), including IRF1 (lower), a known target gene of Phospho-Stat3 (see additional figure containing ChIP-qPCR data).
Chromatin Immunoprecipitation Image 2: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 17

Cell Signaling Technology Logo
Chromatin immunoprecipitations were performed with cross-linked chromatin from Hep G2 cells starved overnight and treated with IL-6 (100 ng/ml) for 30 minutes, and either Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody or of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using human IRF-1 promoter primers, SimpleChIP® Human c-Fos Promoter Primers #4663, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
Chromatin Immunoprecipitation Image 3: Phospho-Stat3 (Tyr705) (D3A7) Rabbit Monoclonal Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.