Human/Mouse Smad2/3 Antibody
R&D Systems, part of Bio-Techne | Catalog # AF3797
Key Product Details
Validated by
Biological Validation
Species Reactivity
Validated:
Human, Mouse
Cited:
Human, Mouse, Rat, Frog - Xenopus (African Clawed Frog)
Applications
Validated:
Chromatin Immunoprecipitation (ChIP), Immunocytochemistry, Immunohistochemistry, Simple Western, Western Blot
Cited:
Chromatin Immunoprecipitation (ChIP), Flow Cytometry, Immunocytochemistry, Immunohistochemistry, Immunoprecipitation, Simple Western, Western Blot
Label
Unconjugated
Antibody Source
Polyclonal Goat IgG
Product Specifications
Immunogen
E. coli-derived recombinant human Smad3
Ser2-Ala230
Accession # P84022
Ser2-Ala230
Accession # P84022
Specificity
Detects human, mouse, and rat Smad2/3 in Western blots. Predicted to detect rat based on sequence homology.
Clonality
Polyclonal
Host
Goat
Isotype
IgG
Scientific Data Images for Human/Mouse Smad2/3 Antibody
Detection of Human/Mouse Smad2/3 by Western Blot.
Western blot shows lysates of A549 human lung carcinoma cell line, Jurkat human acute T cell leukemia cell line, HEK293 human embryonic kidney cell line, and C2C12 mouse myoblast cell line. PVDF membrane was probed with 0.5 µg/mL Goat Anti-Human/Mouse Smad2/3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3797) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). For additional reference, recombinant human Smad2 and Smad3 were included. Specific bands for Smad2 were detected at approximately 64 and 58 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 2.
Detection of Smad2/3-regulated Genes by Chromatin Immunoprecipitation.
Jurkat human acute T cell leukemia cell line treated with 50 ng/mL PMA and 200 ng/mL calcium ionomycin for 30 minutes was fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. Smad2/3/DNA complexes were immunoprecipitated using 5 µg Goat Anti-Human/Mouse Smad2/3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3797) or control antibody (Catalog # AB-108-C) for 15 minutes in an ultrasonic bath, followed by Biotinylated Anti-Goat IgG Secondary Antibody (Catalog # BAF109). Immunocomplexes were captured using 50 µL of MagCellect Streptavidin Ferrofluid (Catalog # MAG999) and DNA was purified using chelating resin solution. Thec-mycpromoter was detected by standard PCR.
Smad2/3 in MCF‑7 Human Cell Line.
Smad2/3 was detected in immersion fixed MCF-7 human breast cancer cell line induced (upper panel) or non-induced (lower panel) to undergo epithelial-mesenchymal transition (EMT) using Goat Anti-Human/Mouse Smad2/3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3797) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; Catalog # NL001) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm and, in EMT-induced cells, nuclei. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
Applications for Human/Mouse Smad2/3 Antibody
Application
Recommended Usage
Chromatin Immunoprecipitation (ChIP)
5 µg/5 x 106 cells
Sample: PMA and calcium ionomycin treated Jurkat human acute T cell leukemia cell line chromatin, c-myc promoter detected by standard PCR
Sample: PMA and calcium ionomycin treated Jurkat human acute T cell leukemia cell line chromatin, c-myc promoter detected by standard PCR
Immunocytochemistry
5-15 µg/mL
Sample: Immersion fixed MCF‑7 human breast cancer cell line non-induced or induced to undergo epithelial-mesenchymal transition
Sample: Immersion fixed MCF‑7 human breast cancer cell line non-induced or induced to undergo epithelial-mesenchymal transition
Immunohistochemistry
3-15 µg/mL
Sample: Immersion fixed paraffin-embedded sections of human brain (cortex and cerebellum)
Sample: Immersion fixed paraffin-embedded sections of human brain (cortex and cerebellum)
Simple Western
25 µg/mL
Sample: A549 human lung carcinoma cell line, COLO 205 human colorectal adenocarcinoma cell line, and HT‑29 human colon adenocarcinoma cell line
Sample: A549 human lung carcinoma cell line, COLO 205 human colorectal adenocarcinoma cell line, and HT‑29 human colon adenocarcinoma cell line
Western Blot
0.5 µg/mL
Sample: A549 human lung carcinoma cell line, Jurkat human acute T cell leukemia cell line, HEK293 human embryonic kidney cell line, and C2C12 mouse myoblast cell line
Sample: A549 human lung carcinoma cell line, Jurkat human acute T cell leukemia cell line, HEK293 human embryonic kidney cell line, and C2C12 mouse myoblast cell line
Please Note: Optimal dilutions of this antibody should be experimentally determined.
Reviewed Applications
Read 3 reviews rated 4.7 using AF3797 in the following applications:
Formulation, Preparation, and Storage
Purification
Antigen Affinity-purified
Reconstitution
Reconstitute at 0.2 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.
Formulation
Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
Shipping
Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
- 12 months from date of receipt, -20 to -70 °C as supplied.
- 1 month, 2 to 8 °C under sterile conditions after reconstitution.
- 6 months, -20 to -70 °C under sterile conditions after reconstitution.
Background: Smad2/3
Smads are a family of intracellular proteins that transmit transforming growth factor beta (TGF-beta) superfamily signals from the cell surface to the nucleus. The Smad family is divided into three subclasses: receptor regulated Smads, (Smads 1, 2, 3, 5 and 8); the common partner, (Smad4); and the inhibitory Smads, (Smads 6 and 7). The binding of TGF-beta or activin to their cognate receptor induces phosphorylation of Smads 2 and 3. The activated Smads associate with the common-mediator subunit, Smad4, and the heteromeric complex translocates into the nucleus to initiate transcription. Smad3, also known as Mothers Against Decapentaplegic homolog 3 (MADH3), shares 83% amino acid identity with Smad2, also known as Mothers Against Decapentaplegic homolog 2 (MADH2). Human Smad2 has 99% identity to mouse and rat Smad2. Human Smad3 has 99% identity to mouse and rat Smad3.
Long Name
Mothers Against DPP Homolog 2/3
Alternate Names
hMAD-2, hSMAD2, JV18, JV18-1SMAD, mothers against DPP homolog 2 (Drosophila), MAD homolog 2, Mad protein homolog, Mad, mothers against decapentaplegic homolog 2, MADH2MGC34440, MADR2MGC22139, Mad-related protein 2, mother against DPP homolog 2, mothers against decapentaplegic homolog 2, Mothers against DPP homolog 2, Sma- and Mad-related protein 2, SMAD 2, SMAD family member 2MAD, mothers against decapentaplegic homolog 2 (Drosophila), SMAD, mothers against DPP homolog 2, Smad2
UniProt
Additional Smad2/3 Products
Product Documents for Human/Mouse Smad2/3 Antibody
Product Specific Notices for Human/Mouse Smad2/3 Antibody
For research use only
Loading...
Loading...
Loading...
Loading...