Cell Signaling Technology

Product Pathways - TGF-beta/Smad Signaling

Phospho-Smad3 (Ser423/425) (C25A9) Rabbit mAb #9520

smad   smad3  

No. Size Price
9520S 100 µl ( 10 western blots ) ¥3,960.00 现货查询 购买询价
9520T 20 µl ( 2 western blots ) ¥1,500.00 现货查询 购买询价
9520 carrier free & custom formulation / quantityemail request
Applications Dilution Species-Reactivity Sensitivity MW (kDa) Isotype
W 1:1000 Human,Mouse,Rat, Endogenous 52 Rabbit IgG
IP 1:50
ChIP 1:100

Species cross-reactivity is determined by western blot.

Applications Key: W=Western Blotting, IP=Immunoprecipitation, ChIP=Chromatin IP,


Species predicted to react based on 100% sequence homology: Monkey, Xenopus, Zebrafish, Bovine,

Specificity / Sensitivity

Phospho-Smad3 (Ser423/425) (C25A9) Rabbit mAb detects endogenous levels of Smad3 when phosphorylated at Ser423/425. This antibody does not cross-react reactivity with other family members.

Phospho-Smad3 (Ser423/425) (C25A9) Rabbit mAb兔单抗可以识别 423/425位丝氨酸磷酸化的内源性Smad3蛋白.本抗体与其他家族成员不发生交叉反应。

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser423/425 of Smad3.


Western Blotting

Western Blotting

Western blot analysis of extracts from HT-1080 cells, untreated or treated with TGF-β1, TGFR inhibitor SB-431542 or BMP-2, using Phospho-Smad3 (Ser423/425) (C25A9) Rabbit mAb (top) or total Smad3 (C67H9) Rabbit mAb #9523 (bottom).

使用Phospho-Smad3 (Ser423/425) (C25A9) Rabbit mAb兔单抗(上)或总Smad3 (C67H9) Rabbit mAb兔单抗 #9523 (下)对未处理或经过 TGF-β1, TGFR 抑制剂 SB-431542 或BMP-2处理的HT-1080细胞提取物进行western blot实验。

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HaCaT cells treated with Human TGF-β3 #3706 (7 ng/ml) for 1 h and either 5 μl of Phospho-Smad3 (Ser423/425) (C25A9) Rabbit mAb or 2 μl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human CDKN1A Intron 1 Primers #4669, SimpleChIP® Human ID1 Promoter Primers #5139, human c-Myc intron 1 primers, 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.

使用SimpleChIP® Enzymatic Chromatin IP Kit (磁珠偶联) #9003对经过人TGF-β3 #3706 (7 ng/ml)处理1小时的4 x 106 HaCaT细胞和5μl Phospho-Smad3 (Ser423/425) (C25A9)兔单抗或2μlNormal Rabbit IgG#2729进行染色质免疫共沉淀。富集到的DNA使用人CDKN1A内含子1引物#4669, SimpleChIP® 人ID1启动子引物#5139,人c-Myc内含子引物和SimpleChIP® 人α Satellite Repeat 引物#4486进行荧光实时PCR定量。各样品沉淀得到的DNA量比对于input染色质总量进行相对定量,Input染色质量设定值为1。

Western Blotting

Western Blotting

Western blot analysis of extracts from HT-1080, C2C12, or KNRK cells, untreated (-) or treated with TGF-β (10 ng/ml, 30 min; +), using Phospho-Smad3 (Ser423/425) (C25A9) Rabbit mAb (upper) or total Smad3 (C67H9) Rabbit mAb #9523 (lower).


Members of the Smad family of signal transduction molecules are components of a critical intracellular pathway that transmit TGF-β signals from the cell surface into the nucleus. Three distinct classes of Smads have been defined: the receptor-regulated Smads (R-Smads), which include Smad1, 2, 3, 5, and 8; the common-mediator Smad (co-Smad), Smad4; and the antagonistic or inhibitory Smads (I-Smads), Smad6 and 7 (1-5). Activated type I receptors associate with specific R-Smads and phosphorylate them on a conserved carboxy terminal SSXS motif. The phosphorylated R-Smad dissociates from the receptor and forms a heteromeric complex with the co-Smad (Smad4), allowing translocation of the complex to the nucleus. Once in the nucleus, Smads can target a variety of DNA binding proteins to regulate transcriptional responses (6-8).

Smad信号转导分子家族成员是胞内通路中TGF-β信号从细胞表面传送到细胞核的重要组成部分。Smad 可被分为三类:受体调控型Smad(简称R-Smad),主要包括Smad1,2,3,5和8;共同介导型Smad(co-Smad)包括Smad4;还有拮抗型或称抑制型Smad(I-Smads),包括Smad6和Smad7[1-5]。激活的I型受体与特异的R-Smad有关,并能在R-Smad的保守的C端SSXS基序处对其磷酸化。磷酸化的R-Smad与受体分离,再与co-Smad(Smad4)形成异侧复合物,使复合物迁移入核。一旦进入核内,Smad可以各种DNA绑定蛋白为目标调控转录反应。[6-8]

Following stimulation by TGF-β, Smad2 and Smad3 become phosphorylated at carboxyl terminal serine residues (Ser465 and 467 on Smad2; Ser423 and 425 on Smad3) by TGF-β Receptor I. Phosphorylated Smad 2/3 can complex with Smad4 and translocate to the nucleus to regulate gene expression (9-11).

TGF-β刺激后,Smad2和Smad3羧基末端丝氨酸残基(Smad2的465位和467位丝氨酸,Smad3的423和425位丝氨酸)被TGF-β Receptor 1磷酸化。磷酸化的Smad2/3与Smad4形成复合物进入细胞核调控基因表达(9-11)。

  1. Heldin, C.H. et al. (1997) Nature 390, 465-71.
  2. Attisano, L. and Wrana, J.L. (1998) Curr Opin Cell Biol 10, 188-94.
  3. Derynck, R. et al. (1998) Cell 95, 737-40.
  4. Massagué, J. (1998) Annu Rev Biochem 67, 753-91.
  5. Whitman, M. (1998) Genes Dev 12, 2445-62.
  6. Wu, G. et al. (2000) Science 287, 92-7.
  7. Attisano, L. and Wrana, J.L. (2002) Science 296, 1646-7.
  8. Moustakas, A. et al. (2001) J Cell Sci 114, 4359-69.
  9. Abdollah, S. et al. (1997) J. Biol. Chem. 272, 27678-27685.
  10. Souchelnytskyi, S. et al. (1997) J. Biol. Chem. 272, 28107-28115.
  11. Liu, X. et al. (1997) Proc. Natl. Acad. Sci. USA 94, 10669-10674.

Application References

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For Research Use Only. Not For Use In Diagnostic Procedures.

U.S. Patent No. 5,675,063.

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

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

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