Revision 6

#12000

Store at +4C

PathScan® Total SMAD2/3 Sandwich ELISA Kit

1 Kit

(96 assays)

Species Cross Reactivity:
H M R Mk Mi

UniProt ID:
#P84022, #Q15796

Entrez-Gene Id:
#4088, #4087

CST Logo
Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Product Includes Product # Quantity Color Storage Temp
SMAD2/3 Mouse mAb Coated Microwells6523596 tests+4C
SMAD2/3 Rabbit Detection mAb139811 eaGreen (Lyophilized)+4C
Anti-rabbit IgG, HRP-linked Antibody (ELISA Formulated)132721 eaRed (Lyophilized)+4C
Detection Antibody Diluent1333911 mlGreen+4C
HRP Diluent1351511 mlRed+4C
TMB Substrate700411 ml+4C
STOP Solution700211 ml+4C
Sealing Tape545032 ea+4C
ELISA Wash Buffer (20X)980125 ml+4C
ELISA Sample Diluent1108325 mlBlue+4C
Cell Lysis Buffer (10X)980315 ml-20C

Kit contents scale proportionally with size, except sealing tape.
Example: The V1 kit contains 5X the listed quantities above, but will exclude the sealing tape.

For the “C” and “V” kits, the supplied 96-well strip plate consists of twelve 8-well strips in a support frame. This enables custom plate configurations.

Description

The PathScan® Total SMAD2/3 Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that recognizes endogenous levels of SMAD2 and SMAD3 proteins. A SMAD2/3 Mouse Antibody has been coated on the microwells. After incubation with cell lysates, SMAD2/3 proteins (phospho and nonphospho) are captured by the coated antibody. Following extensive washing, a SMAD2/3 Rabbit Detection Antibody is added to detect captured SMAD2/3 proteins. Anti-rabbit IgG, HRP-linked Antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of SMAD2 and SMAD3 proteins.

*Antibodies in this kit are custom formulations specific to kit.

Specificity/Sensitivity

PathScan® Total SMAD2/3 Sandwich ELISA Kit recognizes endogenous levels of total SMAD2 and SMAD3 protein in human cells, as shown in Figure 1. This kit sensitivity is shown in Figure 2. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

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).

Trademarks and Patents

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

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

All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 6

Cell Signaling Technology Logo
Figure 1. Treatment of HeLa cells with hTGF-β3 stimulates phosphorylation of SMAD2 at Ser465/467 or SMAD3 at Ser423/425, as detected by PathScan® Phospho-SMAD2 (Ser465/467)/SMAD3 (Ser423/425) Sandwich ELISA Kit #12001, but does not affect the level of total SMAD2 or SMAD3 protein detected by PathScan® Total SMAD2/3 Sandwich ELISA Kit. The absorbance readings at 450 nm are shown in the top figure, while the corresponding western blots using SMAD2/3 (D7G7) XP® Rabbit mAb #8685 (left panel), Phospho-SMAD2 (Ser465/467) (138D4) Rabbit mAb #3108 (center panel), or a phospho-SMAD3 (Ser423/425) Rabbit mAb (right panel) are shown in the bottom figure.
ELISA Image 1: PathScan<sup>®</sup> Total SMAD2/3 Sandwich ELISA Kit
Figure 2. The relationship between the protein concentration of lysates from untreated and TGF-β3-treated HeLa cells and the absorbance at 450 nm as detected by the PathScan® Total SMAD2/3 Sandwich ELISA Kit is shown. Starved HeLa cells (85% confluence) were treated with 10 ng/ml hTGF-β3 for 30 min at 37ºC.
ELISA Image 2: PathScan<sup>®</sup> Total SMAD2/3 Sandwich ELISA Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.