Phospho-Stat3 (Tyr705) Recombinant Rabbit mAb (S-515-360)

Catalog Number: S0B1094 Application: WB, ICC, IP, ChIP Reactivity: Hu Conjugation: Unconjugated Brand: Starter

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$100 USD

Size: 25μl 100μl

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Product Details

Product Specification

Host	Rabbit
Antigen	Phospho-Stat3 (Tyr705)
Synonyms	Signal transducer and activator of transcription 3; Acute-phase response factor; APRF
Immunogen	Synthetic Peptide
Location	Nucleus
Accession	P40763
Clone Number	S-515-360
Antibody Type	Recombinant mAb
Isotype	IgG
Application	WB, ICC, IP, ChIP
Reactivity	Hu
Positive Sample	Jurkat treated with IFN-α, HeLa treated with IFN-α
Predicted Reactivity	Pg, Cw
Purification	Protein A
Concentration	0.5 mg/ml
Conjugation	Unconjugated
Physical Appearance	Liquid
Storage Buffer	PBS, 40% Glycerol, 0.05% BSA, 0.03% Proclin 300
Stability & Storage	12 months from date of receipt / reconstitution, -20 °C as supplied

Dilution

application	dilution	species
Dot Blot	1:1000
WB	1:1000	Hu
IP	1:50	Hu
ICC	1:500	Hu
ChIP	1:20-1:50	Hu

Background

Phospho-Stat3 (Tyr705) refers to the phosphorylated form of Signal Transducer and Activator of Transcription 3 (STAT3) at the tyrosine 705 residue. Phosphorylation of STAT3 at Tyr705 is a critical step in the activation of the STAT3 signaling pathway. This phosphorylation event, often induced by cytokines, growth factors, and oncogenes, leads to the formation of STAT3 dimers, which then translocate to the nucleus to initiate gene transcription. STAT3 is involved in various cellular processes, including cell growth, survival, and immune responses. Phospho-Stat3 (Tyr705) is essential for mediating cellular responses to interleukins, KITLG/SCF, LEP, and other growth factors. In the context of immunity, STAT3 plays a crucial role in regulating the differentiation of naive CD4(+) T-cells into T-helper Th17 or regulatory T-cells (Treg), which is controlled by the phosphorylation at Tyr705. Abnormal activation of STAT3, including constitutive phosphorylation at Tyr705, has been observed in various cancers and is associated with tumor growth, survival, and angiogenesis. In some cases, STAT3 activation is driven by autocrine secretion of interleukin-6 (IL-6), leading to continuous activation of the Jak/Stat pathway.

Picture

Western Blot

WB result of Phospho-Stat3 (Tyr705) Recombinant Rabbit mAb
Primary antibody: Phospho-Stat3 (Tyr705) Recombinant Rabbit mAb at 1/5000 dilution
Lane 1: untreated Jurkat whole cell lysate 20 µg
Lane 2: Jurkat treated with 50 ng/ml IFN-α for 30 minutes whole cell lysate 20 µg
Lane 3: untreated HeLa whole cell lysate 20 µg
Lane 4: HeLa treated with 50 ng/ml IFN-α for 30 minutes whole cell lysate 20 µg
Secondary antibody: Goat Anti-rabbit IgG, (H+L), HRP conjugated at 1/10000 dilution
Predicted MW: 88 kDa
Observed MW: 88 kDa

IP

Phospho-Stat3 (Tyr705) Recombinant Rabbit mAb at 1/50 dilution (1 µg) immunoprecipitating Phospho-Stat3 (Tyr705) in 0.4 mg Jurkat+IFN-α (50 ng/ml, 30 min) whole cell lysate.
Western blot was performed on the immunoprecipitate using Phospho-Stat3 (Tyr705) Recombinant Rabbit mAb at 1/1000 dilution.
Secondary antibody (HRP) for IP was used at 1/1000 dilution.
Lane 1: Jurkat+IFN-α (50 ng/ml, 30 min) whole cell lysate 40 µg (Input)
Lane 2: Phospho-Stat3 (Tyr705) Recombinant Rabbit mAb IP in Jurkat+IFN-α (50 ng/ml, 30 min)whole cell lysate
Lane 3: Rabbit monoclonal IgG IP in Jurkat+IFN-α (50 ng/ml, 30 min) whole cell lysate
Predicted MW: 88 kDa
Observed MW: 88 kDa
Exposure time: 90 s

Dot Blot

Dot blot result of Phospho-Stat3 (Tyr705) Recombinant Rabbit mAb
Lane1: Stat3 (Tyr705) phospho peptide
Lane2: Stat3 unmodified peptide
Primary antibody: Phospho-Stat3 (Tyr705) Recombinant Rabbit mAb at 1/1000 dilution
Secondary antibody: Goat Anti-rabbit IgG, (H+L), HRP conjugated at 1/10000 dilution

Immunocytochemistry

ICC analysis of HeLa cells treated with IFN-α (50 ng/ml, 30 min)(top panel) and untreated HeLa cells (below panel). Anti- Phospho-Stat3 (Tyr705) antibody was used at 1/500 dilution (Green) and incubated overnight at 4°C. Goat polyclonal Antibody to Rabbit IgG - H&L (Alexa Fluor® 488) was used as secondary antibody at 1/1000 dilution. The cells were fixed with 100% ice-cold methanol and permeabilized with 0.1% PBS-Triton X-100. Nuclei were counterstained with DAPI (Blue). Counterstain with tubulin (Red).

ChIP

Chromatin immunoprecipitation (ChIP) was performed on HeLa + IFN-α (50 ng/ml, 30 min) cells cross - linked with 1% formaldehyde for 10 min, then chromatin was fragmented by sonication.
Parallel reactions used Phospho-Stat3 (Tyr705) Recombinant Rabbit mAb (S-515-360) and Rabbit mAb IgG Isotype Control (SDT-R173) at 1:50 for immunoprecipitation.
Post - immunoprecipitation, both samples were washed, eluted, and cross - links reversed. Purified DNA was analyzed by qPCR.

qPCR showed the enrichment of c-FOS, IRF1 and SAT-α in Phospho-Stat3 (Tyr705)
Recombinant Rabbit mAb (S-515-360)-immunoprecipitated sample.