Revision 4

#5678Store at -20C

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, IP, IF-IC, FC-FP, ChIP

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

52, 60

SOURCE:

Rabbit

UniProt ID:

#P84022, #Q15796

Entrez-Gene Id:

4088, 4087

Product Information

Product Usage Information

For optimal ChIP results, use 10 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:50
Immunofluorescence (Immunocytochemistry) 1:100
Flow Cytometry (Fixed/Permeabilized) 1:50 - 1:200
Chromatin IP 1:50

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

SMAD2/3 Antibody recognizes endogenous levels of total SMAD2/3 protein and overexpressed SMAD2/3 protein.

Species Reactivity:

Human, Mouse, Rat, Monkey

Species predicted to react based on 100% sequence homology

Xenopus

Source / Purification

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ala208 of human SMAD2 protein. Antibodies are purified by protein A and peptide affinity chromatography.

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).

  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.

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 IP: Immunoprecipitation IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized) ChIP: Chromatin IP

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
#5678

SMAD2/3 Antibody

Western Blotting Image 1: SMAD2/3 Antibody Expand Image
使用 SMAD2/3 Antibody 对来自 ACHN 细胞和 HeLa 细胞的提取物进行蛋白质印迹分析。
No image available
Immunofluorescence Image 1: SMAD2/3 Antibody Expand Image
使用 SMAD2/SMAD3 Antibody(绿色)对血清饥饿(左图)、经人 TGF-β1 #8915(1 μg/mL,30 分钟;中图)处理或经人 TGF-β1 与 SB43152(10 μg/mL,30 分钟;右图)处理的 HT-1080 细胞进行共聚焦免疫荧光分析。使用 DY-554 phalloidin(红色)标记肌动蛋白纤丝。
Flow Cytometry Image 1: SMAD2/3 Antibody Expand Image
与浓度匹配的 Rabbit (DA1E) mAb IgG XP® Isotype Control #3900(虚线)比较,使用 SMAD2/3 Antibody(实线)对 Hela 细胞进行流式细胞分析。Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor®488Conjugate) #4412 用作二抗。
Chromatin Immunoprecipitation Image 1: SMAD2/3 Antibody Expand Image
使用 SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003,以经人 TGF-β3 #3706 (7 ng/ml) 处理 1 小时的 HaCat 细胞中的交联染色质和 SMAD2/3 Antibody 或 Normal Rabbit IgG #2729 进行染色质免疫沉淀分析。使用 SimpleChIP® Human CDKN1A Intron 1 Primers #4669、SimpleChIP® Human ID1 Promoter Primers #5139 和 SimpleChIP® Human α Satellite Repeat Primers #4486,通过实时 PCR 对富集的 DNA 进行定量分析。将每份样品中免疫沉淀的 DNA 的量表现为相对于所输入染色质总量(等于 1)的信号。