AB247460

Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free

RabMAb
Recombinant
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(1 Publication)

Rabbit Recombinant Monoclonal SMAD5 phospho S463 + S465 antibody. Carrier free. Suitable for WB and reacts with Human, Mouse, Rat samples. Cited in 1 publication.

View Alternative Names

MADH5, SMAD5, SMAD family member 5, SMAD 5, hSmad5, JV5-1, Mothers against decapentaplegic homolog 5, MAD homolog 5, Mothers against DPP homolog 5, BSP1, MADH1, MADR1, SMAD1, SMAD family member 1, SMAD 1, hSMAD1, JV4-1, Mad-related protein 1, Mothers against decapentaplegic homolog 1, Transforming growth factor-beta-signaling protein 1, MAD homolog 1, Mothers against DPP homolog 1, BSP-1

4 Images

Lab

Western blot - Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free (AB247460)

This data was developed using ab76296, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

All lanes:

Western blot - Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] (<a href='/en-us/products/primary-antibodies/smad1-smad5-smad9-phospho-s463-s465-s467-antibody-ep7282ay-ab76296'>ab76296</a>) at 1/1000 dilution

Lane 1:

Untreated HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 15 µg

Lane 2:

HeLa treated with 20ng/ml TGF beta 1 for 15 min whole cell lysate at 15 µg

Lane 3:

HeLa treated with 20ng/ml TGF beta 1 for 15 min whole cell lysate, then the membrane treated with Alkaline Phosphatase for 1 hour at 15 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 52 kDa

false

Exposure time: 20s

Unknown

Western blot - Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free (AB247460)

This data was developed using ab76296, the same antibody clone in a different buffer formulation.

All lanes:

Lane 1:

HeLa cell lysate - untreated at 10 µg

Lane 2:

HeLa cell lysate - treated with BMP-4 at 10 µg

Secondary

All lanes:

goat anti-rabbit HRP at 1/2000 dilution

Observed band size: 52 kDa

false

Lab

Western blot - Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free (AB247460)

This data was developed using ab76296, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer : 5% NFDM/TBST

All lanes:

Lane 1:

Untreated NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate at 15 µg

Lane 2:

NIH/3T3 treated with 20ng/ml TGF beta 1 and 50µM MG-132 for 15 min whole cell lysate at 15 µg

Lane 3:

NIH/3T3 treated with 20ng/ml TGF beta 1 and 50µM MG-132 for 15 min whole cell lysate, then the membrane treated with Alkaline Phosphatase for 1 hour at 15 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 52 kDa

false

Exposure time: 100s

Lab

Western blot - Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free (AB247460)

This data was developed using ab76296, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

All lanes:

Lane 1:

Untreated C6 (Rat glial tumor glial cell) whole cell lysate at 15 µg

Lane 2:

C6 treated with 10ng/ml TGF beta 3 and 50µM MG-132 for 30 min whole cell lysate at 15 µg

Lane 3:

C6 treated with 10ng/ml TGF beta 3 and 50µM MG-132 for 30 min whole cell lysate, then the membrane treated with Alkaline Phosphatase for 1 hour at 15 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 52 kDa

false

Exposure time: 7s

Unconjugated

Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY]

Key facts

Host species

Rabbit

Clonality

Monoclonal

Clone number

EP728(2)AY

Isotype

IgG

Carrier free

Yes

Reacts with

Human, Rat, Mouse

Applications

applications

Immunogen

The exact immunogen used to generate this antibody is proprietary information.

Specificity

Detects SMAD5 phosphorylated at Serine 463 and 465. This antibody may cross-react with Smad1 Phospho (pS463/465) and Smad9 Phospho (pS465/467).

Stimulation may be required to allow detection of the phosphorylated protein. Please see images below for recommended treatment conditions and positive controls.

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Reactivity data

{ "title": "Reactivity Data", "filters": { "stats": ["", "Species", "Dilution Info", "Notes"], "tabs": { "all-applications": {"fullname" : "All Applications", "shortname": "All Applications"}, "ICCIF" : {"fullname" : "Immunocytochemistry/ Immunofluorescence", "shortname":"ICC/IF"}, "IP" : {"fullname" : "Immunoprecipitation", "shortname":"IP"}, "FlowCyt" : {"fullname" : "Flow Cytometry", "shortname":"Flow Cyt"}, "WB" : {"fullname" : "Western blot", "shortname":"WB"}, "IHCP" : {"fullname" : "Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)", "shortname":"IHC-P"} }, "product-promise": { "all": "all", "testedAndGuaranteed": "tested", "guaranteed": "expected", "predicted": "predicted", "notRecommended": "not-recommended" } }, "values": { "Human": { "ICCIF-species-checked": "notRecommended", "ICCIF-species-dilution-info": "", "ICCIF-species-notes": "", "IP-species-checked": "notRecommended", "IP-species-dilution-info": "", "IP-species-notes": "", "FlowCyt-species-checked": "notRecommended", "FlowCyt-species-dilution-info": "", "FlowCyt-species-notes": "", "WB-species-checked": "testedAndGuaranteed", "WB-species-dilution-info": "", "WB-species-notes": "", "IHCP-species-checked": "notRecommended", "IHCP-species-dilution-info": "", "IHCP-species-notes": "" }, "Mouse": { "ICCIF-species-checked": "notRecommended", "ICCIF-species-dilution-info": "", "ICCIF-species-notes": "", "IP-species-checked": "notRecommended", "IP-species-dilution-info": "", "IP-species-notes": "", "FlowCyt-species-checked": "notRecommended", "FlowCyt-species-dilution-info": "", "FlowCyt-species-notes": "", "WB-species-checked": "testedAndGuaranteed", "WB-species-dilution-info": "", "WB-species-notes": "", "IHCP-species-checked": "notRecommended", "IHCP-species-dilution-info": "", "IHCP-species-notes": "" }, "Rat": { "ICCIF-species-checked": "notRecommended", "ICCIF-species-dilution-info": "", "ICCIF-species-notes": "", "IP-species-checked": "notRecommended", "IP-species-dilution-info": "", "IP-species-notes": "", "FlowCyt-species-checked": "notRecommended", "FlowCyt-species-dilution-info": "", "FlowCyt-species-notes": "", "WB-species-checked": "testedAndGuaranteed", "WB-species-dilution-info": "", "WB-species-notes": "", "IHCP-species-checked": "notRecommended", "IHCP-species-dilution-info": "", "IHCP-species-notes": "" } } }

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Product details

ab247460 is the carrier-free version of ab76296.

Species reactivity
Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species.
Please contact us for more information.

Patented technology
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

What are the advantages of a recombinant monoclonal antibody?
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 batch production

For more information, read more on recombinant antibodies.

Conjugation ready
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with 1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

Compatibility
This product is compatible with the Maxpar^® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar^® is a trademark of Fluidigm Canada Inc.

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Properties and storage information

Form

Liquid

Purification technique

Affinity purification

Storage buffer

pH: 7.2 - 7.4 Constituents: PBS

Shipped at conditions

Blue Ice

Appropriate short-term storage conditions

+4°C

Appropriate long-term storage conditions

+4°C

Storage information

Do Not Freeze

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Supplementary information

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

The Smad1 SMAD5 and SMAD9 proteins sometimes known as receptor-regulated SMADs (R-SMADs) play significant roles in cellular signaling. These proteins act as intracellular mediators for the transforming growth factor-beta (TGF-β) superfamily of cytokines particularly the bone morphogenetic proteins (BMPs). These proteins are expressed in various tissues including bone heart lung and brain where they facilitate specific receptor signaling. These SMAD proteins typically have molecular masses ranging from about 50 kDa to 60 kDa which varies depending on specific post-translational modifications such as phosphorylation.

Biological function summary

The Smad1 SMAD5 and SMAD9 proteins are transcription factors involved in the regulation of gene expression. Upon activation by BMP receptors these proteins undergo phosphorylation then form complexes with SMAD4 another member of the SMAD family. This heteromeric complex translocates into the nucleus where it influences the expression of genes associated with cell proliferation differentiation and apoptosis. The Smad1 SMAD5 and SMAD9 proteins therefore coordinate diverse cellular processes by modulating transcriptional responses to extracellular signals in developmental and homeostatic contexts.

Pathways

Smad1 SMAD5 and SMAD9 proteins integrate into the BMP signaling pathway which is fundamental in bone and cartilage development. These proteins interact closely with BMP type I receptors through phosphorylation-mediated mechanisms. Another pathway involving these SMAD proteins is the TGF-β pathway where their activity converges with other signaling molecules like SMAD2 and SMAD3 resulting in complex regulatory outcomes in cellular contexts. The interactions within these pathways help mediate processes like embryogenesis and tissue homeostasis illustrating the interplay of cell signaling and transcriptional regulation.

Smad1 SMAD5 and SMAD9 proteins associate with conditions like pulmonary hypertension and cancer. Dysregulation within the BMP signaling pathway mediated by improper functioning of these R-SMADs links to abnormal vascular remodeling and cancerous cell growth due to altered proliferative signaling. Additionally Smad1 and SMAD5 show connections with proteins such as BMPR2 in the context of pulmonary arterial hypertension further emphasizing their role as mediators of disease-related signaling pathways. Addressing these proteins' malfunction provides potential therapeutic intervention points for complex pathological states.

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Product protocols

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

Visit the General protocols
Visit the Troubleshooting

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Target data

Transcriptional regulator that plays a role in various cellular processes including embryonic development, cell differentiation, angiogenesis and tissue homeostasis (PubMed : 12064918, PubMed : 16516194). Upon BMP ligand binding to their receptors at the cell surface, is phosphorylated by activated type I BMP receptors (BMPRIs) and associates with SMAD4 to form a heteromeric complex which translocates into the nucleus acting as transcription factor (PubMed : 9442019). In turn, the hetero-trimeric complex recognizes cis-regulatory elements containing Smad Binding Elements (SBEs) to modulate the outcome of the signaling network (PubMed : 33510867). Non-phosphorylated SMAD5 has a cytoplasmic role in energy metabolism regulation by promoting mitochondrial respiration and glycolysis in response to cytoplasmic pH changes (PubMed : 28675158). Mechanistically, interacts with hexokinase 1/HK1 and thereby accelerates glycolysis (PubMed : 28675158).

See full target information SMAD5 pS463 + S465

Additional targets

SMAD1 phospho S463 + S465

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Publications (1)

Recent publications for all applications. Explore the full list and refine your search

Nature genetics 57:1142-1154 PubMed40229600

2025

Longitudinal single-cell multiomic atlas of high-risk neuroblastoma reveals chemotherapy-induced tumor microenvironment rewiring.

Applications

Unspecified application

Species

Unspecified reactive species

Wenbao Yu,Rumeysa Biyik-Sit,Yasin Uzun,Chia-Hui Chen,Anusha Thadi,Jonathan H Sussman,Minxing Pang,Chi-Yun Wu,Liron D Grossmann,Peng Gao,David W Wu,Aliza Yousey,Mei Zhang,Christina S Turn,Zhan Zhang,Shovik Bandyopadhyay,Jeffrey Huang,Tasleema Patel,Changya Chen,Daniel Martinez,Lea F Surrey,Michael D Hogarty,Kathrin Bernt,Nancy R Zhang,John M Maris,Kai Tan

View all publications

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Product promise

We are committed to supporting your work with high-quality reagents, and we're here for you every step of the way. In the unlikely event that one of our products does not perform as expected, you're protected by our Product Promise.

For full details, please see our Terms & Conditions

Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.

For licensing inquiries, please contact partnerships@abcam.com

Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free

Western blot - Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free (AB247460)

Western blot - Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free (AB247460)

Western blot - Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free (AB247460)

Western blot - Anti-SMAD1 + SMAD5 + SMAD9 (phospho S463 + S465 + S467) antibody [EP728(2)AY] - BSA and Azide free (AB247460)

Related conjugates and formulations (1)

Key facts

Host species

Clonality

Clone number

Isotype

Carrier free

Reacts with

Applications

Immunogen

Specificity

Complementary Products

Primary Antibodies

Secondary Antibodies

Primary Antibodies

Primary Antibodies

Reactivity data

Product details

Properties and storage information

Form

Purification technique

Storage buffer

Shipped at conditions

Appropriate short-term storage conditions

Appropriate long-term storage conditions

Storage information

Supplementary information

Biological function summary

Pathways

Product protocols

Target data

Additional targets

Publications (1)

Product promise