Revision 4
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, W-S

REACTIVITY:

H M R Mi

SENSITIVITY:

Endogenous

MW (kDa):

60

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q15796

Entrez-Gene Id:

4087

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Simple Western™ 1:50 - 1:250

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.

Specificity / Sensitivity

Phospho-SMAD2 (Ser465/467) (138D4) Rabbit mAb detects endogenous levels of SMAD2 only when dually phosphorylated at serines 465 and 467, and may detect SMAD3 phosphorylated at the equivalent sites. This antibody does not cross-react with other SMAD-related proteins.

Species Reactivity:

Human, Mouse, Rat, Mink

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser465/467 of human SMAD2.

Background

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 9; 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-SMADs dissociate from the receptor and form a heteromeric complex with SMAD4, initiating translocation of the heteromeric SMAD complex to the nucleus. Once in the nucleus, SMADs recruit a variety of DNA binding proteins that function to regulate transcriptional activity (6-8).

Following stimulation by TGF-β, Smad2 and Smad3 become phosphorylated at their carboxy-termini (Ser465/467 on Smad2; Ser423/425 on Smad3) by the receptor kinase TGF-β R1 (9-11). Following phosphorylation, Smad2 and Smad3 form a heteromeric complex with the co-Smad family member Smad4. These complexes are translocated to the nucleus where they bind DNA and regulate gene transcription.

  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. Wrana, J.L. (2000) Sci STKE 2000, re1.
  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, 27678-85.
  10. Souchelnytskyi, S. et al. (1997) J Biol Chem 272, 28107-15.
  11. Liu, X. et al. (1997) Proc Natl Acad Sci U S A 94, 10669-74.

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

WB: Western Blotting W-S: Simple Western™

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

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.

限制使用

除非 CST 的合法授书代表以书面形式书行明确同意,否书以下条款适用于 CST、其关书方或分书商提供的书品。 任何书充本条款或与本条款不同的客书条款和条件,除非书 CST 的合法授书代表以书面形式书独接受, 否书均被拒书,并且无效。

专品专有“专供研究使用”的专专或专似的专专声明, 且未专得美国食品和专品管理局或其他外国或国内专管机专专专任何用途的批准、准专或专可。客专不得将任何专品用于任何专断或治专目的, 或以任何不符合专专声明的方式使用专品。CST 专售或专可的专品提供专作专最专用专的客专,且专用于研专用途。将专品用于专断、专防或治专目的, 或专专售(专独或作专专成)或其他商专目的而专专专品,均需要 CST 的专独专可。客专:(a) 不得专独或与其他材料专合向任何第三方出售、专可、 出借、捐专或以其他方式专专或提供任何专品,或使用专品制造任何商专专品,(b) 不得复制、修改、逆向工程、反专专、 反专专专品或以其他方式专专专专专品的基专专专或技专,或使用专品开专任何与 CST 的专品或服专专争的专品或服专, (c) 不得更改或专除专品上的任何商专、商品名称、徽专、专利或版专声明或专专,(d) 只能根据 CST 的专品专售条款和任何适用文档使用专品, (e) 专遵守客专与专品一起使用的任何第三方专品或服专的任何专可、服专条款或专似专专

Revision 4
#3108

Phospho-SMAD2 (Ser465/467) (138D4) Rabbit mAb

Western Blotting Image 1: Phospho-SMAD2 (Ser465/467) (138D4) Rabbit mAb Expand Image
使用 Phospho-SMAD2 (Ser465/467) (138D4) Rabbit mAb(上图)或 SMAD2 Antibody #3102(下图),对未经处理或经 TGF-β 处理的 HeLa 细胞和 NIH/3T3 细胞的提取物进行蛋白质印迹法分析。
Western Blotting Image 1: Phospho-SMAD2 (Ser465/467) (138D4) Rabbit mAb Expand Image
使用 Phospho-SMAD2 (Ser465/467) (138D4) Rabbit mAb #3108 对经 hTGF-β3(10 ng/mL,30 分钟)处理的血清饥饿 HT-1080 细胞的裂解物 (1.0 mg/mL) 进行 Simple Western™ 分析。虚拟泳道式图像(左图)显示一抗稀释比例为 1:50 和 1:250 时的单一靶标条带(如图所示)。对应的电泳图(右图)为一抗稀释比例在 1:50(蓝线)和 1:250(绿线)时沿毛细血管内分子量的化学发光结果。在还原条件下,使用 12-230 kDa 分离模块在 ProteinSimple(BioTechne 品牌)的 Jess™ Simple Western 仪器上进行该实验。