Recombinant Anti-Smad4 antibody [EP618Y] (ab40759)
![Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-288647-anti-smad4-antibody-ep618y-western-blot.jpg "Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)")
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EP618Y] to Smad4
- Suitable for: WB, IHC-P
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-Smad4 antibody [EP618Y]
See all Smad4 primary antibodies -
Description
Rabbit monoclonal [EP618Y] to Smad4 -
Host species
Rabbit -
Tested applications
Suitable for: WB, IHC-Pmore details
Unsuitable for: Flow Cyt,ICC/IF or IP -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
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Positive control
- WB: Wild type HAP1 whole cell lysate; HepG2, Jurkat, NIH/3T3, PC-12, Ramos, C6 and SH-SY5Y cell lysates; Mouse embryo, skin and lung tissue lysates; Human skin, lung and artery tissue lysates. IHC-P: Human lung carcinoma and breast carcinoma tissues.
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General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 40% Glycerol, 0.05% BSA, 59% PBS -
Concentration information loading... -
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EP618Y -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Recombinant Protein
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab40759 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Abreviews | Notes |
|---|---|---|
| WB | (3) |
1/5000. Detects a band of approximately 60 kDa (predicted molecular weight: 65 kDa).
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| IHC-P | (4) |
1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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| Notes |
|---|
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WB
1/5000. Detects a band of approximately 60 kDa (predicted molecular weight: 65 kDa). |
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IHC-P
1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. |
Target
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Function
Common SMAD (co-SMAD) is the coactivator and mediator of signal transduction by TGF-beta (transforming growth factor). Component of the heterotrimeric SMAD2/SMAD3-SMAD4 complex that forms in the nucleus and is required for the TGF-mediated signaling. Promotes binding of the SMAD2/SMAD4/FAST-1 complex to DNA and provides an activation function required for SMAD1 or SMAD2 to stimulate transcription. Component of the multimeric SMAD3/SMAD4/JUN/FOS complex which forms at the AP1 promoter site; required for syngernistic transcriptional activity in response to TGF-beta. May act as a tumor suppressor. -
Involvement in disease
Defects in SMAD4 are a cause of pancreatic cancer (PNCA) [MIM:260350].
Defects in SMAD4 are a cause of juvenile polyposis syndrome (JPS) [MIM:174900]; also known as juvenile intestinal polyposis (JIP). JPS is an autosomal dominant gastrointestinal hamartomatous polyposis syndrome in which patients are at risk for developing gastrointestinal cancers. The lesions are typified by a smooth histological appearance, predominant stroma, cystic spaces and lack of a smooth muscle core. Multiple juvenile polyps usually occur in a number of Mendelian disorders. Sometimes, these polyps occur without associated features as in JPS; here, polyps tend to occur in the large bowel and are associated with an increased risk of colon and other gastrointestinal cancers.
Defects in SMAD4 are a cause of juvenile polyposis/hereditary hemorrhagic telangiectasia syndrome (JP/HHT) [MIM:175050]. JP/HHT syndrome phenotype consists of the coexistence of juvenile polyposis (JIP) and hereditary hemorrhagic telangiectasia (HHT) [MIM:187300] in a single individual. JIP and HHT are autosomal dominant disorders with distinct and non-overlapping clinical features. The former, an inherited gastrointestinal malignancy predisposition, is caused by mutations in SMAD4 or BMPR1A, and the latter is a vascular malformation disorder caused by mutations in ENG or ACVRL1. All four genes encode proteins involved in the transforming-growth-factor-signaling pathway. Although there are reports of patients and families with phenotypes of both disorders combined, the genetic etiology of this association is unknown.
Defects in SMAD4 may be a cause of colorectal cancer (CRC) [MIM:114500]. -
Sequence similarities
Belongs to the dwarfin/SMAD family.
Contains 1 MH1 (MAD homology 1) domain.
Contains 1 MH2 (MAD homology 2) domain. -
Domain
The MH1 domain is required for DNA binding.
The MH2 domain is required for both homomeric and heteromeric interactions and for transcriptional regulation. Sufficient for nuclear import. -
Post-translational
modificationsMonoubiquitinated on Lys-519 by E3 ubiquitin-protein ligase TRIM33. Monoubiquitination hampers its ability to form a stable complex with activated SMAD2/3 resulting in inhibition of TGF-beta/BMP signaling cascade. Deubiqitination by USP9X restores its competence to mediate TGF-beta signaling. -
Cellular localization
Cytoplasm. Nucleus. Cytoplasmic in the absence of ligand. Migrates to the nucleus when complexed with R-SMAD. - Information by UniProt
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Database links
- Entrez Gene: 4089 Human
- Entrez Gene: 17128 Mouse
- Entrez Gene: 50554 Rat
- Omim: 600993 Human
- SwissProt: Q13485 Human
- SwissProt: P97471 Mouse
- SwissProt: O70437 Rat
- Unigene: 75862 Human
see all -
Alternative names
- (Small) Mothers Against Decapentaplegic antibody
- Deleted in Pancreatic Carcinoma 4 antibody
- Deleted in Pancreatic Carcinoma antibody
see all
Images
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Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)
Lane 1: Wild type HAP1 whole cell lysate (20 µg)
Lane 2: SMAD4 knockout HAP1 whole cell lysate (20 µg)
Lane 3: HepG2 whole cell lysate (20 µg)
Lane 4: Jurkat whole cell lysate (20 µg)Lanes 1 - 4: Merged signal (red and green). Green - ab40759 observed at 60 kDa. Red - loading control, ab9484, observed at 37 kDa.
ab40759 was shown to specifically react with SMAD4 in wild type HAP1 cells. No band was observed when SMAD4 knockout HAP1 samples were used. Wild-type and SMAD4 knockout samples were subjected to SDS-PAGE. Ab40759 and ab9484 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
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![Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-250033-anti-smad4-antibody-ep618y-western-blot.jpg)
Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)All lanes : Anti-Smad4 antibody [EP618Y] (ab40759) at 1/5000 dilution (purified)
Lane 1 : SH-SY5Y (Human neuroblastoma cell line from bone marrow) cell lysate
Lane 2 : Ramos (Human Burkitt's lymphoma cell line) cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution
Predicted band size: 65 kDa
Observed band size: 60 kDa why is the actual band size different from the predicted?Blocking and diluting buffer: 5% NFDM/TBST.
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![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-250069-anti-smad4-antibody-ep618y-immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smad4 antibody [EP618Y] (ab40759)Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast carcinoma tissue labelling Smad4 with purified ab40759 at 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
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![Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-254438-anti-smad4-antibody-ep618y-western-blot.jpg)
Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)All lanes : Anti-Smad4 antibody [EP618Y] (ab40759) at 1/5000 dilution
Lane 1 : NIH/3T3 (Mouse embyro fibroblast cell line) cell lysate
Lane 2 : Mouse embryo tissue lysate
Lane 3 : Mouse skin tissue lysate
Lane 4 : Mouse lung tissue lysate
Lane 5 : PC-12 (Rat adrenal gland pheochromocytoma cell line) cell lysate
Lane 6 : C6 cell lysate
Lane 7 : Rat skin tissue lysate
Lane 8 : Rat lung tissue lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 65 kDa
Observed band size: 60 kDa why is the actual band size different from the predicted?
Exposure time: 3 minutesBlocking and dilution buffer: 5% NFDM/TBST.
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![Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-254437-anti-smad4-antibody-ep618y-western-blot.jpg)
Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)All lanes : Anti-Smad4 antibody [EP618Y] (ab40759) at 1/5000 dilution
Lane 1 : SW480 (Human colorectal adenocarcinoma cell line) cell lysate
Lane 2 : HepG2 (Human liver hepatocellular carcinoma cell line) cell lysate
Lane 3 : Jurkat (Human T cell leukemia cell line from peripheral blood) cell lysate
Lane 4 : Human skin tissue lysate
Lane 5 : Human lung tissue lysate
Lane 6 : Human artery tissue lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 65 kDa
Observed band size: 60 kDa why is the actual band size different from the predicted?
Exposure time: 30 secondsBlocking and dilution buffer: 5% NFDM/TBST.
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![Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-18-anti-smad4-antibody-ep618y-westernblot-ramos-human.jpg)
Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)Different batches of ab40759 were tested on Ramos (Human Burkitt's lymphoma B lymphocyte) lysate at 0.7 µg/ml. 15 µg of lysate was loaded in each lane. Bands observed at 60 kDa. -
![Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-250034-anti-smad4-antibody-ep618y-western-blot.jpg)
Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)Anti-Smad4 antibody [EP618Y] (ab40759) at 1/10000 dilution (purified) + NIH/3T3 (Mouse embyro fibroblast cell line) cell lysate at 20 µg
Secondary
Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution
Predicted band size: 65 kDa
Observed band size: 60 kDa why is the actual band size different from the predicted?Blocking and diluting buffer: 5% NFDM/TBST.
-
![Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-250035-anti-smad4-antibody-ep618y-western-blot.jpg)
Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)Anti-Smad4 antibody [EP618Y] (ab40759) at 1/5000 dilution (purified) + PC-12 (Rat adrenal gland pheochromocytoma cell line) cell lysate at 10 µg
Secondary
Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution
Predicted band size: 65 kDa
Observed band size: 60 kDa why is the actual band size different from the predicted?Blocking and diluting buffer: 5% NFDM/TBST.
-
![Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-53870-anti-smad4-antibody-ep618y-western-blot.jpg)
Western blot - Anti-Smad4 antibody [EP618Y] (ab40759)Anti-Smad4 antibody [EP618Y] (ab40759) at 1/5000 dilution (unpurified) + SHSY5Y (Human neuroblastoma cell line from bone marrow) cell lysate at 10 µg
Predicted band size: 65 kDa -
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-53869-anti-smad4-antibody-ep618y-immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smad4 antibody [EP618Y] (ab40759)Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human lung carcinoma tissue labelling Smad4 with unpurified ab40759 at a 1/100 dilution.
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-53866-anti-smad4-antibody-ep618y-immunohistochemistry.jpg)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Smad4 antibody [EP618Y] (ab40759)This image is courtesy of an anonymous AbreviewUnpurified ab40759 staining Smad4 in rat femur tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA for 20 minutes at 22°C; antigen retrieval was by heat mediation in a citrate buffer pH6.0. Samples were incubated with primary antibody (1/200 in blocking buffer) for 2 hours at 20°C. An undiluted HRP-conjugated goat anti-rabbit IgG polyclonal (1/250) was used as the secondary antibody.
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![Anti-Smad4 antibody [EP618Y] (ab40759)](https://www.abcam.com/ps/products/40/ab40759/Images/ab40759-17-benefits-of-recombinant-antibodies.png)
Anti-Smad4 antibody [EP618Y] (ab40759)
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (136)
ab40759 has been referenced in 136 publications.
- Long C et al. FOXO3 is targeted by miR-223-3p and promotes osteogenic differentiation of bone marrow mesenchymal stem cells by enhancing autophagy. Hum Cell 34:14-27 (2021). PubMed: 32920731
- Zhao S et al. Effects of Smad4 on the expression of caspase-3 and Bcl-2 in human gingival fibroblasts cultured on 3D PLGA scaffolds induced by compressive force. Int J Mol Med 47:N/A (2021). PubMed: 33495811
- Zhang T et al. Proteomics reveals the function reverse of MPSSS-treated prostate cancer-associated fibroblasts to suppress PC-3 cell viability via the FoxO pathway. Cancer Med 10:2509-2522 (2021). PubMed: 33704935
- Wang W et al. Positive feedback regulation between USP15 and ERK2 inhibits osteoarthritis progression through TGF-ß/SMAD2 signaling. Arthritis Res Ther 23:84 (2021). PubMed: 33726807
- Xu R et al. MicroRNA-181a promotes epithelial-mesenchymal transition in esophageal squamous cell carcinoma via the TGF-ß/Smad pathway. Mol Med Rep 23:N/A (2021). PubMed: 33760133