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SMAD3 (pS423/S425) ELISA Kit is a single-wash 90-min Simplestep used to quantify SMAD3 (pS423/S425) with a sensitivity of 10 µg/mL. The assay uses a simple mix-wash-read protocol with just one incubation and one wash step.

- Colorimetric Sandwich ELISA - 450 nm readout : works on any standard plate reader
- Cited in over 5 citations

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Images

Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (AB186038), expandable thumbnail
  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (AB186038), expandable thumbnail
  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (AB186038), expandable thumbnail
  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (AB186038), expandable thumbnail
  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (AB186038), expandable thumbnail

Publications

Key facts

Detection method
Colorimetric
Sample types
Tissue Homogenate, Cell Lysate
Assay type
Semi-quantitative
Reactive species
Mouse, Human
Range
10 - 500 µg/mL
Assay time
1h 30m
Sensitivity
= 10 µg/mL

Reactivity data

Application
ELISA
Reactivity
Reacts
Dilution info
-
Notes

-

Target data

Function

SMAD3 is a receptor-regulated SMAD that functions as an intracellular signal transducer and transcriptional modulator, activated by TGF-beta and activin type 1 receptor kinases. It binds to the TRE element in promoters of numerous genes regulated by TGF-beta, and upon forming a complex with SMAD4, activates transcription. Additionally, SMAD3 can form a complex with SMAD4, JUN, and FOS at the AP-1/SMAD site to regulate TGF-beta-mediated transcription. SMAD3 may inhibit wound healing by modulating the growth and migration of primary keratinocytes and altering TGF-beta-mediated monocyte chemotaxis, with this effect being potentially hormone-sensitive. Furthermore, SMAD3 is involved in regulating chondrogenesis and osteogenesis and may inhibit early bone fracture healing. It also positively regulates PDPK1 kinase activity by promoting its dissociation from the 14-3-3 protein YWHAQ, which negatively regulates it. This supplementary information is collated from multiple sources and compiled automatically.

Alternative names

What's included?

1 x 96 Tests
Components
10X Wash Buffer PT
1 x 15 mL
50X Cell Extraction Enhancer Solution
1 x 1 mL
5X Cell Extraction Buffer PTR
1 x 12 mL
Lyophilized SMAD3 (pS423/S425) Control Lysate
1 x 1 Vial
Plate Seal
1 x 1 Unit
SMAD3 (pS423/S425) Capture Antibody
1 x 3 mL
SMAD3 (pS423/S425) Detector Antibody
1 x 3 mL
SimpleStep Pre-Coated 96-Well Microplate (ab206978)
1 x 1 Unit
Stop Solution
1 x 12 mL
TMB Substrate
1 x 12 mL

Recommended products

SMAD3 (pS423/S425) ELISA Kit is a single-wash 90-min Simplestep used to quantify SMAD3 (pS423/S425) with a sensitivity of 10 µg/mL. The assay uses a simple mix-wash-read protocol with just one incubation and one wash step.

- Colorimetric Sandwich ELISA - 450 nm readout : works on any standard plate reader
- Cited in over 5 citations

Key facts

Detection method
Colorimetric
Sample types
Tissue Homogenate, Cell Lysate
Assay type
Semi-quantitative
Reactive species
Mouse, Human
Range
10 - 500 µg/mL
Assay time
1h 30m
Assay Platform
Microplate
Sensitivity
= 10 µg/mL

Precision

Intra assay

Sample
A431 Extract
n
6
C.V.
2.4

Inter assay

Sample
A431
n
3
C.V.
7.4

Storage

Shipped at conditions
Blue Ice
Appropriate short-term storage conditions
+4°C
Appropriate long-term storage conditions
+4°C
Storage information
+4°C

Notes

Abcam's SMAD3 (pS423/S425) in vitro SimpleStep ELISA™ (Enzyme-Linked Immunosorbent Assay) kit is designed for the semi-quantitative measurement of SMAD3 (pS423/S425) protein in human and mouse cells.

The SimpleStep ELISA™ employs a labeled capture and detector antibody which immunocaptures the sample analyte in solution. This entire complex (capture antibody/protein/detector antibody) is in turn immobilized in the well by immunoaffinity via the anti-tag antibody. Samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material; the TMB substrate is then added. The reaction is stopped by addition of Stop Solution which stops the color development and completes any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.

ASSAY SPECIFICITY

The SMAD3 (pS423/S425) assay detects endogenous levels of SMAD3 (GenBank Accession NP_001138574) in cellular lysates, only when phosphorylated at Ser423/425. Based on sequence similarity, cross reaction to SMAD2 may occur.

SPECIES REACTIVITY

This kit detects SMAD3 (pS423/S425) in Human and mouse cell culture extracts. Detection in rat samples is also expected. Other species should be tested on a case-by-case basis.

Serum and plasma samples have not been tested with this kit.

Abcam has not and does not intend to apply for the REACH Authorisation of customers' uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.
Activity summary

Smad3 also known as Mothers against decapentaplegic homolog 3 is a protein that plays a mechanical role in signal transduction. It acts mainly as a transcription factor and gets activated through phosphorylation. The molecular weight of Smad3 is approximately 48 kDa. It is expressed widely across numerous tissues including the cellular nucleus where it executes its function after activation.

Biological function summary

Smad3 acts as a mediator of signal transduction for the TGF-beta (transforming growth factor-beta) superfamily forming a complex with phosphorylated Smad2. This enables it to regulate transcriptional activity influencing cell proliferation differentiation and apoptosis. Smad3 also participates in various cellular processes by interacting with other co-factors and regulatory proteins that aid in fine-tuning its function.

Pathways

Smad3 plays an important role in the TGF-beta signaling pathway where it works closely with Smad4 to propagate the signal. Upon phosphorylation it forms a complex with co-Smad (Smad4) and moves into the nucleus to influence gene expression. Smad3 is also involved in pathways related to oncogenesis and tissue fibrosis indicating its significant role in cellular regulation and response mechanisms.

Associated diseases and disorders

Smad3 is associated with fibrotic diseases and cancers particularly in tissues such as the liver and lungs. Altered Smad3 signaling contributes to the pathological process occurring in fibrotic disorders often interacting with Smad4 in these abnormalities. Dysregulated Smad3 expression or mutations can also lead to oncogenic transformations highlighting its critical involvement in disease states.

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6 product images

  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038), expandable thumbnail

    Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038)

    SMAD3 (pS423/S425) control lysate dilution series in 1X Cell Extraction Buffer PTR.

    Raw data values are shown in the table.

  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038), expandable thumbnail

    Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038)

    Linearity of dilution in representative sample matrices. Cellular lysates were prepared at 3 concentrations in common media containing 1X Cell Extraction Buffer PTR. Data from duplicate measurements of SMAD3 (pS423/S425) are normalized and plotted.

  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038), expandable thumbnail

    Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038)

    SMAD3 (pS423/S425) phosphorylation in response to TGF-beta treatment.

    Induction of SMAD3 (pS423/S425) phosphorylation in HeLa cells in response to TGF-β treatment. HeLa cells were cultured in 96-well tissue culture plates, serum-starved and treated (60 min) with a dose-range of TGF-β before cell lysis. Data from quadruplicate measurements of SMAD3 (pS423/S425) are plotted.

  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038), expandable thumbnail

    Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038)

    SMAD3 (pS423/S425) phosphorylation in response to SB431542 treatment.

    Inhibition of SMAD3 (pS423/S425) phosphorylation in HeLa cells in response to SB431542 treatment. HeLa cells were cultured in 96-well tissue culture plates and treated with a dose-range of SB431542 (60 min). Cells were then stimulated with TGF-β (60 min) and lysed. Data from quadruplicate measurements of SMAD3 (pS423/S425) are plotted.

  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038), expandable thumbnail

    Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038)

    Example of a typical SMAD3 (pS423/S425) control lysate dilution series in 1X Cell Extraction Buffer PTR. The SMAD3 lysate dilution series was prepared as described. Raw data values are shown in the table.

  • Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038), expandable thumbnail

    Sandwich ELISA - SMAD3 (pS423/S425) ELISA Kit (ab186038)

    Example of a typical SMAD3 (pS423/S425) control lysate dilution series in 1X Cell Extraction Buffer PTR.

    The SMAD3 lysate dilution series was prepared as described. Raw data values are shown in the table.

    Kit Control lysates are provided at a concentration that give consistent signal between different lots. Lysates are produced and formulated by signal intensity to be consistent to within 30% of the previous lot. As such, Control lysates are not provided with a protein concentration.

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Product protocols

For this product, it's our understanding that no specific protocols are required. You can:

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