Overview

  • Product name

  • Description

    Rabbit polyclonal to SMAD1/5/9
  • Host species

    Rabbit
  • Tested applications

    Suitable for: ICC/IF, WB, IHC-Pmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Dog, Human, Xenopus laevis, Zebrafish
    Predicted to work with: Rabbit, Horse, Chicken, Guinea pig, Cow, Cat, Pig, Chimpanzee, Drosophila melanogaster
  • Immunogen

    Synthetic peptide within Human SMAD1/5/9 (C terminal). The exact sequence is proprietary.
    Sequence:

    FVKGWGAEYHRQDVTSTPCWIEIHLHGPLQWLDKVLTQMGSPHNPISSVS


    Database link: Q15797

  • Positive control

    • Jurkat cell lysate

Properties

Applications

Our Abpromise guarantee covers the use of ab66737 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
ICC/IF Use a concentration of 1 µg/ml.
WB Use a concentration of 0.625 µg/ml. Detects a band of approximately 62 kDa (predicted molecular weight: 52 kDa). Good results were obtained when blocked with 5% non-fat dry milk in 0.05% PBS-T.
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

Target

  • Relevance

    SMADs are members of the MAD-related family of molecules. MAD-related proteins are a family of intracellular proteins that are essential components in the signaling pathways of the serine/threonine kinase receptors of the transforming growth factor beta superfamily. SMADs can be divided into receptor-regulated SMADs, common-mediator SMAD and inhibitory SMADs. SMAD1, SMAD5 and SMAD9 have high degrees of homology. SMAD8 and SMAD9 are used as alternate names for the same protein. Human SMAD1 is a 465 amino acid protein, human SMAD5 is a 465 amino acid and human SMAD9 is a 430 amino acid protein.
  • Cellular localization

    Cytoplasmic and Nuclear
  • Database links

  • Alternative names

    • AI528653 antibody
    • BSP1 antibody
    • DKFZp781C1895 antibody
    • DKFZp781O1323 antibody
    • Dwarfin A antibody
    • Dwf A antibody
    • Dwfc antibody
    • hSMAD1 antibody
    • hSmad5 antibody
    • JV4 1 antibody
    • JV41 antibody
    • JV5 1 antibody
    • MAD (mothers against decapentaplegic, Drosophila) homolog 1 antibody
    • MAD (mothers against decapentaplegic, Drosophila) homolog 9 antibody
    • MAD homolog 1 antibody
    • MAD homolog 9 antibody
    • MAD homolog1 (mothers against decapentaplegic, Drosophila) antibody
    • Mad related protein 1 antibody
    • MAD, mothers against decapentaplegic homolog 9 antibody
    • Madh1 antibody
    • MADH5 antibody
    • MADH6 antibody
    • MADH9 antibody
    • MADR1 antibody
    • Mlp1 antibody
    • Mothers against decapentaplegic, drosophila, homolog of, 9 antibody
    • Mothers against DPP homolog 1 antibody
    • Mothers against DPP homolog 9 antibody
    • MusMLP antibody
    • OTTHUMP00000178565 antibody
    • OTTMUSP00000022970 antibody
    • OTTMUSP00000022971 antibody
    • OTTMUSP00000022972 antibody
    • OTTMUSP0000002324 antibody
    • OTTMUSP00000023428 antibody
    • Sma- and Mad-related protein 1 antibody
    • SMAD antibody
    • SMAD family member 5 antibody
    • SMAD8 antibody
    • SMAD8A antibody
    • SMAD8B antibody
    • TGF beta signaling protein 1 antibody
    • Transforming growth factor-beta signaling protein 1 antibody
    see all

Images

  • Anti-SMAD1/5/9 antibody (ab66737) at 0.625 µg/ml + Jurkat cell lysate at 10 µg

    Secondary
    HRP conjugated anti-Rabbit IgG at 1/50000 dilution

    Predicted band size: 52 kDa
    Observed band size: 62 kDa
    why is the actual band size different from the predicted?

  • IHC image of ab66737 staining in human heart formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab66737, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • ICC/IF image of ab66737 stained HeLa cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab66737, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

References

This product has been referenced in:

  • Zhang Y  et al. ClC-7 Regulates the Pattern and Early Development of Craniofacial Bone and Tooth. Theranostics 9:1387-1400 (2019). Read more (PubMed: 30867839) »
  • Miao N  et al. Loss of Fam20c causes defects in the acinar and duct structure of salivary glands in mice. Int J Mol Med 43:2103-2117 (2019). Read more (PubMed: 30864688) »
See all 12 Publications for this product

Customer reviews and Q&As

1-7 of 7 Abreviews or Q&A

Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Bone marrow derived mast cells)
Permeabilization
Yes - 0.1% Triton-X 100
Specification
Bone marrow derived mast cells
Blocking step
BSA as blocking agent for 45 minute(s) · Concentration: 5% · Temperature: 4°C
Fixative
Paraformaldehyde

Mrs. Nicole Corder

Verified customer

Submitted Aug 13 2018

Application
Western blot
Loading amount
20 µg
Gel Running Conditions
Reduced Denaturing (10% gel)
Sample
Fruit fly (Drosophila melanogaster) Tissue lysate - whole (testis, ovary and heads)
Specification
testis, ovary and heads
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C

Abcam user community

Verified customer

Submitted Nov 08 2013

Question
Answer

We carry products which recognize mouse SMAD1/9, SMAD2/3 or SMAD1/5/9. There is no antibody currently on our catalog which recognizes all forms of SMAD in mouse. I have added links to our anti- SMAD1/9, SMAD2/3 and SMAD1/5/9 antibodies for your review below: Anti- SMAD1/9; AB108965: https://www.abcam.com/index.html?datasheet=108965 Anti- SMAD2/3; AB65847: https://www.abcam.com/index.html?datasheet=65847 Anti- SMAD1/5/9; AB6673 7: https://www.abcam.com/index.html?datasheet=66737

Read More

Answer

Thank you for contacting us.

Your credit note ID is xxx.

I am sorry that this antibody had a mistake onthe datasheet - which we have corrected in the meantime. I have asked our accounting department to issue a credit note for you, which can be redeemed against the invoice of a future order by passing it on to your purchasing department. To avoid confusion, please ensure your accounts department is aware of how the credit note is being used. If you have questions on how to use the credit note, please contact our accounting department.

Our accounting department can be contacted by email at us.credits@abcam.com or by telephone using the information at the Contact Us link in the top right corner of our website. Please refer to the credit note ID in any correspondence with our accounting department.

The credit note ID is for your reference only and does not automatically guarantee the credit.

I hope this experience will not prevent you from purchasing other products from us in the future. Our Scientific Support team is always at your service, should you require further expert advice.

Read More

Answer

Thank you for contacting us.

After BLASTing the immunizing peptide sequence of ab66737 , there is a 100% homology to SMAD1 and SMAD9 and a 93% homology to SMAD5. I would expect this antibody to recognize and react with all three of these SMAD family members.We will update or datasheet on this occasion

However, the ab75273 has not been tested against the Smad9 recombinant protein, but as the full length protein has been used as an immunogen, there may be some cross-reactivity observed against Smad9 as well.

Read More

Answer

Thank you for contacting us.

Ab75273 can detect smad1 and smad5. It’s validated with each recombinant protein.

The molecular weight of smad1 and smad5 is same and that’s the reason why the predicted band size is same.

The possibility of detecting sharing region of smad1 and smad5 is very high because their sequence alignment is 89%.

However, we cannot officially comment which region it detects because it’s not mapped the epitope this antibody detect.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Read More
Application
Western blot
Sample
Human Cell lysate - whole cell (Calu6 cell line)
Loading amount
25 µg
Specification
Calu6 cell line
Gel Running Conditions
Reduced Denaturing (8%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C

Abcam user community

Verified customer

Submitted Jul 21 2010

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