Recombinant
RabMAb

Recombinant Anti-Smad1 (phospho S463 + S465) antibody [EPR20662-20] - BSA and Azide free (ab228457)

Overview

  • Product name

    Anti-Smad1 (phospho S463 + S465) antibody [EPR20662-20] - BSA and Azide free
    See all Smad1 primary antibodies
  • Description

    Rabbit monoclonal [EPR20662-20] to Smad1 (phospho S463 + S465) - BSA and Azide free
  • Host species

    Rabbit
  • Specificity

    Based on sequence homology this antibody also reacts with Smad5 (phospho S463/S465) and Smad9 (phospho S465/S467).
  • Tested applications

    Suitable for: IP, WB, ICC/IF, Dot blotmore details
  • Species reactivity

    Reacts with: Mouse, Human
  • Immunogen

    Synthetic peptide within Human Smad1 aa 450-550 (phospho S463 + S465). The exact sequence is proprietary.
    Database link: Q15797

  • Positive control

    • ICC/IF: NIH3T3 cells FBS-deprived overnight before treatment with 50 ng/ml hBMP2 for 30min.
  • General notes

    Ab228457 is the carrier-free version of ab226821. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

     

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    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.

    ab228457 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

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

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab228457 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
IP Use at an assay dependent concentration.
WB Use at an assay dependent concentration. Detects a band of approximately 60 kDa (predicted molecular weight: 52 kDa).
ICC/IF Use at an assay dependent concentration.
Dot blot Use at an assay dependent concentration.

Target

  • Function

    Transcriptional modulator activated by BMP (bone morphogenetic proteins) type 1 receptor kinase. SMAD1 is a receptor-regulated SMAD (R-SMAD). SMAD1/OAZ1/PSMB4 complex mediates the degradation of the CREBBP/EP300 repressor SNIP1.
  • Tissue specificity

    Ubiquitous. Highest expression seen in the heart and skeletal muscle.
  • 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 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. Co-localizes with LEMD3 at the nucleus inner membrane.
  • Information by UniProt
  • Database links

  • Alternative names

    • BSP-1 antibody
    • BSP1 antibody
    • HsMAD1 antibody
    • JV4-1 antibody
    • JV41 antibody
    • MAD homolog 1 antibody
    • MAD mothers against decapentaplegic homolog 1 antibody
    • Mad related protein 1 antibody
    • Mad-related protein 1 antibody
    • MADH1 antibody
    • MADR1 antibody
    • Mothers against decapentaplegic homolog 1 antibody
    • Mothers against DPP homolog 1 antibody
    • SMA- AND MAD-RELATED PROTEIN 1 antibody
    • SMAD 1 antibody
    • SMAD family member 1 antibody
    • SMAD mothers against DPP homolog 1 antibody
    • Smad1 antibody
    • SMAD1_HUMAN antibody
    • TGF beta signaling protein 1 antibody
    • Transforming growth factor-beta-signaling protein 1 antibody
    see all

Images

  • Dot blot analysis of Smad1 (phospho S463 + S465) labeled with ab226821 at 1/1000 dilution.

    Lane 1: Smad1 (phospho S463/S465) peptide;

    Lane 2: Smad1 (phospho S463) peptide;

    Lane 3: Smad1 (phospho S465) peptide;

    Lane 4: Smad1 peptide (not phosphorylated);

    Lane 5: Smad5 (phospho S463/S465) peptide;

    Lane 6: Smad5 (phospho S463) peptide;

    Lane 7: Smad5 (phospho S465) peptide;

    Lane 9: Smad5 peptide (not phosphorylated).

    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution was used as secondary antibody.

    Blocking/Dilution buffer: 5% NFDM/TBST.

    Exposure time: 3 minutes.

    Based on sequence homology, this antibody cross-reacts with Smad5 (phospho S463/S465) and Smad9 (phospho S465/S467).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab226821).

  • Smad 1 (phospho S463 + S465) was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryo fibroblast cell line) grown in serum-free media overnight, then treated with 50 ng/ml BMP2 for 30 minutes, whole cell lysate with ab226821 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab226821 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

    Lane 1: NIH/3T3 grown in serum-free media overnight, then treated with 50 ng/ml BMP2 for 30 minutes, whole cell lysate 10 μg (Input).

    Lane 2: ab226821 IP in NIH/3T3 grown in serum-free media overnight, then treated with 50 ng/ml BMP2 for 30 minutes, whole cell lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab226821 in NIH/3T3 grown in serum-free media overnight, then treated with 50 ng/ml BMP2 for 30 minutes, whole cell lysate.

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

    Exposure time: 10 seconds.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab226821).

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embryo fibroblast cell line) cells labeling Smad1 (phospho S463 + S465) with ab226821 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Nuclear staining in hBMP2-treated NIH/3T3 cells. Cells were FBS-deprived overnight before treatment with 50 ng/ml hBMP2 for 30 minutes.

    The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab226821).

References

ab228457 has not yet been referenced specifically in any publications.

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