Anti-Phospho-Smad3 (Ser423 + Ser425) antibody (bs-3425R)

Anti-Phospho-Smad3 (Ser423 + Ser425) antibody

DATASHEET

Host:Rabbit

Target Protein:Phospho-Smad3 (Ser423 + Ser425)

IR:Immunogen Range:CS(p-S)V(p-S)

Clonality:Polyclonal

Isotype:IgG

Entrez Gene:4088

Swiss Prot:P84022

Source:KLH conjugated Synthesised phosphopeptide derived from human Smad3 around the phosphorylation site of Ser423/425:CS(p-S)V(p-S) 

Purification:affinity purified by Protein A

Storage:0.01M TBS(pH7.4) with 1% BSA, 0.03% Proclin300 and 50% Glycerol. Shipped at 4℃. Store at -20 °C for one year. Avoid repeated freeze/thaw cycles.

Background:Smad3 is a 50 kDa member of a family of proteins that act as key mediators of TGF beta superfamily signaling in cell proliferation, differentiation and development. The Smad family is divided into three subclasses: receptor regulated Smads, activin/TGF beta receptor regulated (Smad2 and 3) or BMP receptor regulated (Smad 1, 5, and 8); the common partner, (Smad4) that functions via its interaction to the various Smads; and the inhibitory Smads, (Smad6 and 7). Activated Smad3 oligomerizes with Smad4 upon TGF beta stimulation and translocates as a complex into the nucleus, allowing its binding to DNA and transcription factors. Phosphorylation of the two TGF beta dependent serines 423 and 425 in the C terminus of Smad3 is critical for Smad3 transcriptional activity and TGF beta signaling.

Size:100ul

Concentration:1mg/ml

Applications:WB(1:500-2000)
ELISA(1:5000-10000)
IHC-P(1:100-500)
IHC-F(1:100-500)
Flow-Cyt(1μg/Test)
ICC(1:100-500)
IF(1:100-500)

Cross Reactive Species:Human
Mouse
Rat
Chicken
Dog
Pig
Cow
Horse
.

For research use only. Not intended for diagnostic or therapeutic use.

VALIDATION IMAGES

Sample:
Lane 1: Cerebrum (Mouse) Lysate at 40 ug
Lane 2: Heart (Mouse) Lysate at 40 ug
Lane 3: Testis (Mouse) Lysate at 40 ug
Lane 4: Skin (Mouse) Lysate at 40 ug
Lane 5: Kidney (Mouse) Lysate at 40 ug
Lane 6: Cerebrum (Rat) Lysate at 40 ug
Lane 7: Testis (Rat) Lysate at 40 ug
Lane 8: Skin (Rat) Lysate at 40 ug
Lane 9: Kidney (Rat) Lysate at 40 ug
Lane 10: Huvec (Human) Cell Lysate at 30 ug
Lane 11: A549 (Human) Cell Lysate at 30 ug
Lane 12: Hela (Human) Cell Lysate at 30 ug
Lane 13: HT1080 (Human) Cell Lysate at 30 ug
Lane 14: A431 (Human) Cell Lysate at 30 ug
Primary: Anti-Phospho-Smad3 (Ser423 + Ser425) (bs-3425R) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 52 kD
Observed band size: 54 kD

Sample:
293T (Human) Cell Lysate at 30 ug
Cerebrum (Mouse) Lysate at 40 ug
Cerebrum (Rat) Lysate at 40 ug
Ovary (Rat) Lysate at 40 ug
HT1080 (Human) Cell Lysate at 30 ug
Jurkat(Human) Cell Lysate at 30 ug
Liver (Rat) Lysate at 40 ug
Primary: Anti- Phospho-Smad3 (Ser423 + Ser425) (bs-3425R) at 1/300 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 52 kD
Observed band size: 63 kD

Paraformaldehyde-fixed, paraffin embedded (human gastric carcinoma); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Smad3 (Ser423 + Ser425)) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (rat heart); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Smad3 (Ser423 + Ser425)) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (rat kidney); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Smad3 (Ser423 + Ser425)) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (rat skin); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Smad3 (Ser423 + Ser425)) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Smad3 (Ser423 + Ser425)) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (rat testis); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Smad3 (Ser423 + Ser425)) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (mouse kidney); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Smad3 (Ser423 + Ser425)) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (mouse skin); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Smad3 (Ser423 + Ser425)) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (mouse brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (Phospho-Smad3 (Ser423 + Ser425)) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Tissue/cell: human liver carcinoma; 4% Paraformaldehyde-fixed and paraffin-embedded;
Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min;
Incubation: Anti-Phospho-Smad3(Ser423/425) Polyclonal Antibody, Unconjugated(bs-3425R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining