Phospho-Smad3 (Ser423 + Ser425) Polyclonal Antibody

$Lane 1: mouse embryo lysates Lane 2: mouse brain lysates probed with Anti Phospho-Smad3(Ser423\/425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 in 4˚C. Followed by conjugation to secondary antibody (bs-0295G-HRP) at 1:3000 90min in 37˚C. Predicted band 47kD. Observed band size: 47kD.$

Lane 1: mouse embryo lysates Lane 2: mouse brain lysates probed with Anti Phospho-Smad3(Ser423\/425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 in 4˚C. Followed by conjugation to secondary antibody (bs-0295G-HRP) at 1:3000 90min in 37˚C. Predicted band 47kD. Observed band size: 47kD.

$Formalin-fixed and paraffin embedded human liver carcinoma labeled with Rabbit Anti-Smad3 (Ser423 + Ser425) Polyclonal Antibody (bs-3425R) at 1:200 followed by conjugation to the secondary antibody and DAB staining\\n$

Formalin-fixed and paraffin embedded human liver carcinoma labeled with Rabbit Anti-Smad3 (Ser423 + Ser425) Polyclonal Antibody (bs-3425R) at 1:200 followed by conjugation to the secondary antibody and DAB staining\\n

$HUVEC cells(black) were fixed with 4% PFA for 10min at room temperature,permeabilized with 90% ice-cold methanol for 20 min at -20\u2103, and incubated in 5% BSA blocking buffer for 30 min at room temperature. Cells were then stained with Smad3 (Ser423 + Ser425) Polyclonal Antibody(bs-3425R)at 1:50 dilution in blocking buffer and incubated for 30 min at room temperature, washed twice with 2% BSA in PBS, followed by secondary antibody(blue) incubation for 40 min at room temperature. Acquisitions of 20,000 events were performed. Cells stained with primary antibody (green), and isotype control (orange).$

HUVEC cells(black) were fixed with 4% PFA for 10min at room temperature,permeabilized with 90% ice-cold methanol for 20 min at -20\u2103, and incubated in 5% BSA blocking buffer for 30 min at room temperature. Cells were then stained with Smad3 (Ser423 + Ser425) Polyclonal Antibody(bs-3425R)at 1:50 dilution in blocking buffer and incubated for 30 min at room temperature, washed twice with 2% BSA in PBS, followed by secondary antibody(blue) incubation for 40 min at room temperature. Acquisitions of 20,000 events were performed. Cells stained with primary antibody (green), and isotype control (orange).

$Paraformaldehyde-fixed, paraffin embedded Mouse brain; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.$

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\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.

$Paraformaldehyde-fixed, paraffin embedded Mouse skin; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.$

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\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.

$Paraformaldehyde-fixed, paraffin embedded Mouse kidney; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.$

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\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.

$Paraformaldehyde-fixed, paraffin embedded Rat testis; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.$

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\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.

$Paraformaldehyde-fixed, paraffin embedded Rat brain; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.$

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\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.

$Paraformaldehyde-fixed, paraffin embedded Rat skin; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.$

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\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.

$Paraformaldehyde-fixed, paraffin embedded Rat kidney; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.$

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\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.

$Paraformaldehyde-fixed, paraffin embedded Rat heart; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.$

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\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.

$Paraformaldehyde-fixed, paraffin embedded Human gastric carcinoma; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.$

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\u00b0C for 30min; Antibody incubation with Smad3 (Ser423 + Ser425) Polyclonal Antibody, Unconjugated (bs-3425R) at 1:200 overnight at 4\u00b0C, DAB staining.

Lane 1: Mouse Cerebrum lysates; Lane 2: Mouse Heart lysates; Lane 3: Mouse Testis lysates; Lane 4: Mouse Skin lysates; Lane 5: Mouse Kidney lysates; Lane 6: Rat Cerebrum lysates; Lane 7: Rat Testis lysates; Lane 8: Rat Skin lysates; Lane 9: Rat Kidney lysates; Lane 10: Human HUVEC cell lysates; Lane 11: Human A549 cell lysates; Lane 12: Human Hela cell lysates; Lane 13: Human HT1080 cell lysates; Lane 14: Human A431 cell lysates probed with Smad3 Polyclonal Antibody, Unconjugated (bs-3425R) at 1:1000 dilution and 4˚C overnight incubation. Followed by conjugated secondary antibody incubation at 1:20000 for 60 min at 37˚C.

$Hela cells were fixed with 4% PFA for 10min at room temperature,permeabilized with 90% ice-cold methanol for 20 min at -20\u2103, and incubated in 5% BSA blocking buffer for 30 min at room temperature. Cells were then stained with Smad3 (Ser423 + Ser425) Antibody(bs-3425R)at 1:100 dilution in blocking buffer and incubated for 30 min at room temperature, washed twice with 2%BSA in PBS, followed by secondary antibody incubation for 40 min at room temperature. Acquisitions of 20,000 events were performed. Cells stained with primary antibody (green), and isotype control (orange).$

Hela cells were fixed with 4% PFA for 10min at room temperature,permeabilized with 90% ice-cold methanol for 20 min at -20\u2103, and incubated in 5% BSA blocking buffer for 30 min at room temperature. Cells were then stained with Smad3 (Ser423 + Ser425) Antibody(bs-3425R)at 1:100 dilution in blocking buffer and incubated for 30 min at room temperature, washed twice with 2%BSA in PBS, followed by secondary antibody incubation for 40 min at room temperature. Acquisitions of 20,000 events were performed. Cells stained with primary antibody (green), and isotype control (orange).

Applications

WB
ELISA
FCM
IHC-P
IHC-F
IF(IHC-P)
IF(IHC-F)
IF(ICC)

Reactivity

Human
Mouse
Rat
Pig

Predicted Reactivity

Dog
Cow
Horse
Chicken

Overview
Catalog #	bs-3425R
Product Name	Phospho-Smad3 (Ser423 + Ser425) Polyclonal Antibody
Applications	WB, ELISA, FCM, IHC-P, IHC-F, IF(IHC-P), IF(IHC-F), IF(ICC)
Specificity	These phosphorylation sites are homologous across the species listed.
Reactivity	Human, Mouse, Rat, Pig
Predicted Reactivity	Dog, Cow, Horse, Chicken
Specifications
Conjugation	Unconjugated
Host	Rabbit
Source	KLH conjugated synthetic phosphopeptide derived from human Smad3 around the phosphorylation site of Ser423/425 [CS(p-S)V(p-S)]
Modification Site	Ser423 + Ser425
Clonality	Polyclonal
Clone #	#REF!
Isotype	IgG
Concentration	1ug/ul
Purification	Purified by Protein A.
Storage Buffer	0.01M TBS(pH7.4) with 1% BSA, 0.02% Proclin300 and 50% Glycerol.
Storage Condition	Shipped at 4_. Store at -20_ for one year. Avoid repeated freeze/thaw cycles.
Target
Gene ID	4088
Swiss Prot	P84022
Subcellular location	Cytoplasm, Nucleus
Synonyms	LDS3; LDS1C; MADH3; JV15-2; HSPC193; HsT17436; Mothers against decapentaplegic homolog 3; MAD homolog 3; Mad3; Mothers against DPP homolog 3; hMAD-3; SMAD family member 3; SMAD 3; Smad3; hSMAD3
Background	Receptor-regulated SMAD (R-SMAD) that is an intracellular signal transducer and transcriptional modulator activated by TGF-beta (transforming growth factor) and activin type 1 receptor kinases. Binds the TRE element in the promoter region of many genes that are regulated by TGF-beta and, on formation of the SMAD3/SMAD4 complex, activates transcription. Also can form a SMAD3/SMAD4/JUN/FOS complex at the AP-1/SMAD site to regulate TGF-beta-mediated transcription. Has an inhibitory effect on wound healing probably by modulating both growth and migration of primary keratinocytes and by altering the TGF-mediated chemotaxis of monocytes. This effect on wound healing appears to be hormone-sensitive. Regulator of chondrogenesis and osteogenesis and inhibits early healing of bone fractures. Positively regulates PDPK1 kinase activity by stimulating its dissociation from the 14-3-3 protein YWHAQ which acts as a negative regulator.
Application Dilution
WB	1:300-5000
ELISA	1:500-1000
FCM	1:20-100
IHC-P	1:200-400
IHC-F	1:100-500
IF(IHC-P)	1:50-200
IF(IHC-F)	1:50-200
IF(ICC)	1:50-200