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    products/primary-antibodies/smad5-antibody-ep619y-ab40771.pdf

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Signal Transduction Signaling Pathway Nuclear Signaling SMADs
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RecombinantRabMAb

Recombinant Anti-SMAD5 antibody [EP619Y] (ab40771)

  • Datasheet
  • SDS
Reviews (4)Q&A (1)References (24)

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Western blot - Anti-SMAD5 antibody [EP619Y] (ab40771)
  • Western blot - Anti-SMAD5 antibody [EP619Y] (ab40771)
  • Flow Cytometry (Intracellular) - Anti-SMAD5 antibody [EP619Y] (ab40771)
  • Immunocytochemistry/ Immunofluorescence - Anti-SMAD5 antibody [EP619Y] (ab40771)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD5 antibody [EP619Y] (ab40771)
  • Western blot - Anti-SMAD5 antibody [EP619Y] (ab40771)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD5 antibody [EP619Y] (ab40771)
  • Flow Cytometry (Intracellular) - Anti-SMAD5 antibody [EP619Y] (ab40771)
  • Anti-SMAD5 antibody [EP619Y] (ab40771)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EP619Y] to SMAD5
  • Suitable for: Flow Cyt (Intra), IHC-P, WB, ICC/IF, Dot blot
  • Reacts with: Mouse, Rat, Human, African green monkey

Conjugates logo Related conjugates and formulations

Alexa Fluor® 488 Carrier Free PE

You may also be interested in

Secondary
Product image
Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
Knockout
Human SMAD5 knockout A549 cell line (ab277890)
Protein
Product image
Recombinant Human SMAD5 protein (ab84651)

View more associated products

Overview

  • Product name

    Anti-SMAD5 antibody [EP619Y]
    See all SMAD5 primary antibodies
  • Description

    Rabbit monoclonal [EP619Y] to SMAD5
  • Host species

    Rabbit
  • Tested applications

    Suitable for: Flow Cyt (Intra), IHC-P, WB, ICC/IF, Dot blotmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human, African green monkey
  • Immunogen

    Synthetic peptide within Human SMAD5 aa 200-300. The exact sequence is proprietary.

  • Positive control

    • WB: HEK293 and Cos-1 whole cell lysate. Flow Cyt (intra): PC-12 and HEK293 cells. ICC/IF: HeLa cells. IHC-P: Human testis tissue and human skin tissue.
  • 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
    For more information see here.

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

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

Properties

  • 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: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 40% Glycerol, 0.05% BSA, 59% PBS
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EP619Y
  • Isotype

    IgG
  • Research areas

    • Signal Transduction
    • Signaling Pathway
    • Nuclear Signaling
    • SMADs
    • Epigenetics and Nuclear Signaling
    • Nuclear Signaling Pathways
    • SMADs
    • Stem Cells
    • Signaling Pathways
    • TGF beta
    • Cytoplasmic
    • Developmental Biology
    • Organogenesis
    • Gut development
    • Liver development

Associated products

  • Alternative Versions

    • Alexa Fluor® 488 Anti-SMAD5 antibody [EP619Y] (ab206042)
    • PE Anti-SMAD5 antibody [EP619Y] (ab213662)
    • Anti-SMAD5 antibody [EP619Y] - BSA and Azide free (ab238952)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Recombinant Protein

    • Recombinant Human SMAD5 protein (ab84651)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab40771 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
Flow Cyt (Intra)
1/30 - 1/100.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

IHC-P (1)
1/50. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB (2)
1/1000 - 1/5000. Detects a band of approximately 52 kDa (predicted molecular weight: 52 kDa).
ICC/IF (1)
1/50 - 1/100.
Dot blot
Use at an assay dependent concentration.
Notes
Flow Cyt (Intra)
1/30 - 1/100.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

IHC-P
1/50. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
WB
1/1000 - 1/5000. Detects a band of approximately 52 kDa (predicted molecular weight: 52 kDa).
ICC/IF
1/50 - 1/100.
Dot blot
Use at an assay dependent concentration.

Target

  • Function

    Transcriptional modulator activated by BMP (bone morphogenetic proteins) type 1 receptor kinase. SMAD5 is a receptor-regulated SMAD (R-SMAD).
  • Tissue specificity

    Ubiquitous.
  • Sequence similarities

    Belongs to the dwarfin/SMAD family.
    Contains 1 MH1 (MAD homology 1) domain.
    Contains 1 MH2 (MAD homology 2) domain.
  • Post-translational
    modifications

    Phosphorylated on serine by BMP (bone morphogenetic proteins) type 1 receptor kinase.
    Ubiquitin-mediated proteolysis by SMAD-specific E3 ubiquitin ligase SMURF1.
  • Cellular localization

    Cytoplasm. Nucleus. Cytoplasmic in the absence of ligand. Migrates to the nucleus when complexed with SMAD4.
  • Target information above from: UniProt accession Q99717 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 4090 Human
    • Entrez Gene: 17129 Mouse
    • Entrez Gene: 59328 Rat
    • Omim: 603110 Human
    • SwissProt: Q99717 Human
    • SwissProt: P97454 Mouse
    • SwissProt: Q9R1V3 Rat
    • Unigene: 167700 Human
    • Unigene: 272920 Mouse
    • Unigene: 146857 Rat
    see all
  • Alternative names

    • DKFZp781C1895 antibody
    • DKFZp781O1323 antibody
    • Dwfc antibody
    • hSmad5 antibody
    • JV5 1 antibody
    • JV5-1 antibody
    • MAD homolog 5 antibody
    • MAD, mothers against decapentaplegic homolog 5 antibody
    • MADH 5 antibody
    • MADH5 antibody
    • Mothers against decapentaplegic homolog 5 antibody
    • mothers against decapentaplegic, drosophila, homolog of, 5 antibody
    • Mothers against DPP homolog 5 antibody
    • MusMLP antibody
    • SMA and MAD related protein 5 antibody
    • SMAD 5 antibody
    • SMAD family member 5 antibody
    • SMAD, mothers against DPP homolog 5 antibody
    • Smad5 antibody
    • SMAD5_HUMAN antibody
    see all

Images

  • Western blot - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Western blot - Anti-SMAD5 antibody [EP619Y] (ab40771)
    All lanes : Anti-SMAD5 antibody [EP619Y] (ab40771) at 1/1000 dilution

    Lane 1 : 3T3-L1 (Mouse embryonic fibroblast) lysate
    Lane 2 : Neuro-2a (Mouse neuroblastoma neuroblast)
    Lane 3 : F9 (Mouse embryonal carcinoma epithelial cell) lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

    Predicted band size: 52 kDa
    Observed band size: 52 kDa


    Exposure time: 3 minutes


    Blocking and dilution buffer: 5% NFDM/TBST

  • Western blot - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Western blot - Anti-SMAD5 antibody [EP619Y] (ab40771)
    All lanes : Anti-SMAD5 antibody [EP619Y] (ab40771) at 1/5000 dilution (purified)

    Lane 1 : HEK293 whole cell lysate
    Lane 2 : COS-1 whole cell lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution (HRP goat anti-rabbit IgG (H+L))

    Predicted band size: 52 kDa
    Observed band size: 52 kDa



    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

  • Flow Cytometry (Intracellular) - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Flow Cytometry (Intracellular) - Anti-SMAD5 antibody [EP619Y] (ab40771)

    Overlay histogram showing PC-12 cells fixed in 4% PFA and stained with purified ab40771 at a dilution of 1/100 (red line). The secondary antibody used was Alexa Fluorr® 488 goat anti-rabbit at a dilution of 1/500. Rabbit monoclonal IgG was used as an isotype control (black line) and cells incubated in the absence of both primary and secondary antibody were used as a negative control (blue line). 

     

     

  • Immunocytochemistry/ Immunofluorescence - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Immunocytochemistry/ Immunofluorescence - Anti-SMAD5 antibody [EP619Y] (ab40771)

    Immunofluorescence staining of HeLa cells with purified ab40771 at a working dilution of 1/100, counter-stained with DAPI. The secondary antibody was Alexa Fluor® 488 goat anti-rabbit (ab150077), used at a dilution of 1/1000. ab7291, a mouse anti-tubulin antibody (1/1000), was used to stain tubulin along with ab150120 (Alexa Fluor® 594 goat anti-mouse, 1/1000), shown in the top right hand panel. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative controls are shown in bottom middle and right hand panels - for negative control 1, purified ab40771 was used at a dilution of 1/500 followed by an Alexa Fluor® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500. For negative control 2, ab7291 (mouse anti-tubulin) was used at a dilution of 1/500 followed by an Alexa Fluor® 488 goat anti-rabbit antibody (ab150077) at a dilution of 1/400.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Immunohistochemical staining of paraffin embedded human testis with purified ab40771 at a working dilution of 1/50. The secondary antibody used is ab97051, a goat anti-rabbit IgG (H&L) at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
  • Western blot - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Western blot - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Anti-SMAD5 antibody [EP619Y] (ab40771) at 1/1000 dilution (unpurified) + Cos-1 cell lysate at 10 µg

    Predicted band size: 52 kDa
    Observed band size: 52 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SMAD5 antibody [EP619Y] (ab40771)

    Unpurified ab40771 (4µg/ml) staining SMAD5 in human skin using an automated system (DAKO Autostainer Plus). Using this protocol there is strong staining of nuclear/cytoplasmic compartments within the stratum granulosum.
    Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.

  • Flow Cytometry (Intracellular) - Anti-SMAD5 antibody [EP619Y] (ab40771)
    Flow Cytometry (Intracellular) - Anti-SMAD5 antibody [EP619Y] (ab40771)

    Overlay histogram showing HEK293 cells stained with unpurified ab40771 (red line). The cells were fixed with methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab40771, 1/50 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit monoclonal IgG (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HEK293 cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions. 

     

     

  • Anti-SMAD5 antibody [EP619Y] (ab40771)
    Anti-SMAD5 antibody [EP619Y] (ab40771)

Protocols

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (24)

Publishing research using ab40771? Please let us know so that we can cite the reference in this datasheet.

ab40771 has been referenced in 24 publications.

  • Jin T  et al. CHI3L1 promotes Staphylococcus aureus-induced osteomyelitis by activating p38/MAPK and Smad signaling pathways. Exp Cell Res 403:112596 (2021). PubMed: 33826950
  • Herberg S  et al. Scaffold-free human mesenchymal stem cell construct geometry regulates long bone regeneration. Commun Biol 4:89 (2021). PubMed: 33469154
  • Xiao X  et al. Endothelial Bone Morphogenetic Protein 2 (Bmp2) Knockout Exacerbates Hemochromatosis in Homeostatic Iron Regulator (Hfe) Knockout Mice but not Bmp6 Knockout Mice. Hepatology 72:642-655 (2020). PubMed: 31778583
  • Wang CY  et al. Ablation of Hepatocyte Smad1, Smad5, and Smad8 Causes Severe Tissue Iron Loading and Liver Fibrosis in Mice. Hepatology 70:1986-2002 (2019). PubMed: 31127639
  • Herberg S  et al. Combinatorial morphogenetic and mechanical cues to mimic bone development for defect repair. Sci Adv 5:eaax2476 (2019). PubMed: 31489377
View all Publications for this product

Customer reviews and Q&As

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1-5 of 5 Abreviews or Q&A

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-SMAD5 antibody [EP619Y]

Excellent
Abreviews
Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Rat Tissue sections (Nasal turbinates)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: 10mM citrate pH6
Permeabilization
Yes - 0.1% Triton X-100
Specification
Nasal turbinates
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 20°C
Fixative
Formaldehyde
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Catherine Heyward

Verified customer

Submitted Feb 06 2023

Immunocytochemistry/ Immunofluorescence abreview for Anti-SMAD5 antibody [EP619Y]

Excellent
Abreviews
Abreviews
abreview image
Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (normal human dermal fibroblasts)
Permeabilization
Yes - 0.1% Triton X-100
Specification
normal human dermal fibroblasts
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 20°C
Fixative
Paraformaldehyde
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Catherine Heyward

Verified customer

Submitted Feb 06 2023

Western blot abreview for Anti-SMAD5 antibody [EP619Y]

Good
Abreviews
Abreviews
abreview image
Application
Western blot
Sample
Mouse Cell lysate - whole cell (liver)
Gel Running Conditions
Reduced Denaturing (4-20)
Loading amount
100 µg
Specification
liver
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

Submitted May 01 2015

Western blot abreview for Anti-SMAD5 antibody [EP619Y]

Excellent
Abreviews
Abreviews
abreview image
Application
Western blot
Sample
Human Cell lysate - nuclear (hESCs)
Loading amount
20 µg
Specification
hESCs
Treatment
BMP4 24h
Gel Running Conditions
Reduced Denaturing (4-12%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 4% · Temperature: 23°C
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

Submitted Sep 02 2008

Question

Product code: 40771
Inquiry: J'aimerais connaitre la concentration de l'anticorps Anti-SMAD5 antibody [EP619Y] (ab40771) Merci

Read More

Abcam community

Verified customer

Asked on May 02 2012

Answer

Merci de nous avoir contactés.

La concentration exactedu lot de ab40771 que nous avons actuellement en stock (lot GR7392-3) est : 0.264 mg/ml.



J'espère que cette information vous sera utile. N'hésitez pas à nous contacter de nouveau si vous avez d'autres questions.

Read More

Abcam Scientific Support

Answered on May 02 2012

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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