Smad4 Antibody [Unconjugated]

HAF019). A specific band was detected for Smad4 at approximately 60 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1." > HAF019). A specific band was detected for Smad4 at approximately 60 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1." title="Western blot shows lysates of HeLa human cervical epithelial carcinoma cell line, Jurkat human acute T cell leukemia cell line, and K562 human chronic myelogenous leukemia cell line. PVDF membrane was probed with 0.25 µg/mL of Goat Anti-Human Smad4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2097) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF019). A specific band was detected for Smad4 at approximately 60 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1." />

Western blot shows lysates of HeLa human cervical epithelial carcinoma cell line, Jurkat human acute T cell leukemia cell line, and K562 human chronic myelogenous leukemia cell line. PVDF membrane was probed with 0.25 µg/mL of Goat Anti-Human Smad4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2097) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # <a class=

219-IL) overnight was fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. Smad4/DNA complexes were immunoprecipitated using 5 µg Goat Anti-Human Smad4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2097) 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. Thep21promoter was detected by standard PCR." > 219-IL) overnight was fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. Smad4/DNA complexes were immunoprecipitated using 5 µg Goat Anti-Human Smad4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2097) 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. Thep21promoter was detected by standard PCR." title="Jurkat human acute T cell leukemia cell line treated with 10 ng/mL Recombinant Human IL-12 (Catalog # 219-IL) overnight was fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. Smad4/DNA complexes were immunoprecipitated using 5 µg Goat Anti-Human Smad4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2097) 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. Thep21promoter was detected by standard PCR." />

Jurkat human acute T cell leukemia cell line treated with 10 ng/mL Recombinant Human IL-12 (Catalog # <a class=

NL001) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Non-adherent Cells." > NL001) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Non-adherent Cells." title="Smad4 was detected in immersion fixed human peripheral blood mononuclear cells (PBMCs) using Goat Anti-Human Smad4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2097) at 15 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. View our protocol for Fluorescent ICC Staining of Non-adherent Cells." />

Smad4 was detected in immersion fixed human peripheral blood mononuclear cells (PBMCs) using Goat Anti-Human Smad4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2097) at 15 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; Catalog # <a class=

siRNA knockdown of Smad1, -2 and 4 does not impact RVFV replication.HSAECs transfected with media alone (Mock) or siRNAs (75nM) directed against a nontargeting control (NTC), Smad1, -2, or -4 were infected with MP12 virus (MOI 0.1). Both extracellular media supernatants (A) and protein lysates (B,C) were harvested at 9 and 24hpi. A) Infectious viral titers were determined by plaque assay. Data plotted as a bar graph of the mean with standard deviation of four replicates. Protein lysates from single (B) and double knockdowns (C) were analyzed by western blot for actin, total Smad1, -2, and -4 expression. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/29408900), licensed under a CC-BY license. Not internally tested by R&D Systems.

siRNA knockdown of Smad1, -2 and 4 does not impact RVFV replication.HSAECs transfected with media alone (Mock) or siRNAs (75nM) directed against a nontargeting control (NTC), Smad1, -2, or -4 were infected with MP12 virus (MOI 0.1). Both extracellular media supernatants (A) and protein lysates (B,C) were harvested at 9 and 24hpi. A) Infectious viral titers were determined by plaque assay. Data plotted as a bar graph of the mean with standard deviation of four replicates. Protein lysates from single (B) and double knockdowns (C) were analyzed by western blot for actin, total Smad1, -2, and -4 expression. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/29408900), licensed under a CC-BY license. Not internally tested by R&D Systems.

• Reactivity: Hu
• Applications: WB, ChIP, ICC/IF
• Clonality: Polyclonal
• Host: Goat
• Conjugate: Unconjugated
• Concentration: LYOPH

Smad4 Antibody [Unconjugated] Summary

• Immunogen: E. coli-derived recombinant human Smad4
  Pro139-Asp332
  Accession # Q13485
• Specificity: Detects human Smad4 in direct ELISAs and Western blots. In direct ELISAs and Western blots, approximately 15% cross-reactivity with recombinant human (rh) Smad1 and less than 5% cross-reactivity with rhSmad9, rhSmad5, and rhSmad7 is observed.
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Goat
• Gene: SMAD4
• Purity Statement: Antigen Affinity-purified

Applications/Dilutions

• Dilutions:
  • Chromatin Immunoprecipitation (ChIP) 5 ug/5 x 10^6 cells
  • Immunocytochemistry 5-15 ug/mL
  • Western Blot 0.25 ug/mL

Packaging, Storage & Formulations

• 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.
• Buffer: 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.
• Preservative: No Preservative
• Concentration: LYOPH
• Reconstitution Instructions: Reconstitute at 0.2 mg/mL in sterile PBS.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Smad4 Antibody [Unconjugated]

• DPC4
• DPC4SMAD, mothers against DPP homolog 4 (Drosophila)
• hSMAD4
• JIP
• MAD homolog 4
• MADH4
• MADH4mothers against decapentaplegic homolog 4
• mothers against decapentaplegic homolog 4
• mothers against decapentaplegic, Drosophila, homolog of, 4
• SMAD 4
• SMAD family member 4MAD, mothers against decapentaplegic homolog 4 (Drosophila)
• SMAD, mothers against DPP homolog 4
• Smad4

Background

Smads are a family of intracellular proteins that transmit transforming growth factor beta (TGF-beta ) superfamily signals from the cell surface to the nucleus. Upon signal-induced phosphorylation, Smad subunits associate with the common-mediator subunit, Smad4. This heteromeric complex then translocates into the nucleus to exert transcriptional comodulator activity.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for Smad4 Antibody (AF2097)(9)

Publications using AF2097

• LL Lin, ER Kost, CL Lin, P Valente, CM Wang, MG Kolonin, AC Daquinag, X Tan, N Lucio, CN Hung, CP Wang, NB Kirma, TH Huang PAI-1-Dependent Inactivation of SMAD4-Modulated Junction and Adhesion Complex in Obese Endometrial Cancer Cell Rep, 2020-10-13;33(2):108253. 2020-10-13 [PMID: 33053339] (Immunoprecipitation, ICC, Simple Western, Human)
• Yuan Fang, Xu Han, Jianang Li, Tiantao Kuang, Wenhui Lou HEATR1 Deficiency Promotes Chemoresistance via Upregulating ZNF185 and Downregulating SMAD4 in Pancreatic Cancer Journal of Oncology 2020-05-26 [PMID: 32565799] (Western Blot, Human)
• Cynthia de la Fuente, Chelsea Pinkham, Deemah Dabbagh, Brett Beitzel, Aura Garrison, Gustavo Palacios et al. Phosphoproteomic analysis reveals Smad protein family activation following Rift Valley fever virus infection PLOS ONE 2018-02-06 [PMID: 29408900] (Chromatin Immunoprecipitation (ChIP), Human)
• N Kumeda, Y Ogawa, Y Akimoto, H Kawakami, M Tsujimoto, R Yanoshita Characterization of Membrane Integrity and Morphological Stability of Human Salivary Exosomes Biol. Pharm. Bull., 2017-01-01;40(8):1183-1191. 2017-01-01 [PMID: 28768999] (Western Blot, Human)
• Jie Yang, Ya Wang, Zhen Zeng, Long Qiao, Liang Zhuang, Qinglei Gao et al. Smad4 deletion in blood vessel endothelial cells promotes ovarian cancer metastasis International Journal of Oncology 2017-05-01 [PMID: 28393199] (Immunohistochemistry, Human)
• MJ Moen, HH Adams, JH Brandsma, DH Dekkers, U Akinci, S Karkampoun, M Quevedo, CE Kockx, Z Ozgür, WF van IJcken, J Demmers, RA Poot An interaction network of mental disorder proteins in neural stem cells Transl Psychiatry, 2017-04-04;7(4):e1082. 2017-04-04 [PMID: 28375211] (Immunoprecipitation, Human)
• LM de Kroon, R Narcisi, GG van den Ak, EL Vitters, EN Blaney Dav, GJ van Osch, PM van der Kr SMAD3 and SMAD4 have a more dominant role than SMAD2 in TGF?-induced chondrogenic differentiation of bone marrow-derived mesenchymal stem cells Sci Rep, 2017-02-27;7(0):43164. 2017-02-27 [PMID: 28240243] (Western Blot, Human)
• Kazunobu Isogaya, Daizo Koinuma, Shuichi Tsutsumi, Roy-Akira Saito, Keiji Miyazawa, Hiroyuki Aburatani et al. A Smad3 and TTF-1/NKX2-1 complex regulates Smad4-independent gene expression Cell Research 2014-08-01 [PMID: 25060702]
• Kim SW, Yoon SJ, Chuong E, Oyolu C, Wills AE, Gupta R, Baker J Chromatin and transcriptional signatures for Nodal signaling during endoderm formation in hESCs. Dev. Biol., 2011-06-29;357(2):492-504. 2011-06-29 [PMID: 21741376] (ChIP, Human)

Bioinformatics

• Gene Symbol: SMAD4
• Uniprot:
  • Q13485()