Recombinant
RabMAb

Recombinant Anti-Smad1 (phospho S463 + S465) antibody [EPR20662-29] (ab214423)

Overview

  • Product name

    Anti-Smad1 (phospho S463 + S465) antibody [EPR20662-29]
    See all Smad1 primary antibodies
  • Description

    Rabbit monoclonal [EPR20662-29] to Smad1 (phospho S463 + S465)
  • 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: WB, Dot blot, IPmore details
  • Species reactivity

    Reacts with: Mouse, Human
  • Immunogen

    Recombinant fragment within Human Smad1 aa 450 to the C-terminus. The exact sequence is proprietary.
    Database link: Q15797

  • Positive control

    • Dot Blot: Smad1 (phospho S463/S465) peptide, Smad1 (phospho S465) peptide, Smad5 (phospho S463/S465) peptide, Smad5 (phospho S465) peptide. WB: Calyculin (ab141784) treated HeLa cell lysate; BMP2 treated NIH/3T3 cell lysate. IP: BMP2 treated NIH/3T3 cell lysate.
  • General notes

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

Properties

Applications

Our Abpromise guarantee covers the use of ab214423 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
WB 1/1000. Detects a band of approximately 60 kDa (predicted molecular weight: 52 kDa).
Dot blot 1/1000.
IP 1/30.

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

  • All lanes : Anti-Smad1 (phospho S463 + S465) antibody [EPR20662-29] (ab214423) at 1/1000 dilution

    Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell) grown in serum-free media overnight, whole cell lysate
    Lane 2 : HeLa grown in serum-free media overnight, then treated with 100ng/ml Calyculin A (ab141784) for 15 minutes, Calyculin A was removed, followed by treatment with 100ng/ml BMP2 for 30 minutes, whole cell lysate
    Lane 3 : NIH/3T3 (mouse embryonic fibroblast) grown in serum-free media overnight, whole cell lysate
    Lane 4 : NIH/3T3 grown in serum-free media overnight, then treated with 50ng/ml BMP2 for 30 minutes, whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 52 kDa
    Observed band size: 60 kDa
    why is the actual band size different from the predicted?


    Exposure time: 10 seconds


    Blocking/Dilution: 5% NFDM/TBST.

  • Dot blot analysis of Smad1 (phospho S463/S465) peptide (Lane 1), Smad1 (phospho S463) peptide (Lane 2), Smad1 (phospho S465) peptide (Lane 3), Smad1 non-phospho peptide (Lane 4), Smad5 (phospho S463/S465) peptide (Lane 5), Smad5 (phospho S463) peptide (Lane 6), Smad5 (phospho S465) peptide (Lane 7) and Smad5 non-phospho peptide (Lane 8) using ab214423 at 1/1,000 dilution followed by Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100,000 dilution.

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

    Exposure time: 30 seconds.

  • Smad1 (phospho S463/S465) was immunoprecipitated from 0.35 mg of NIH/3T3 (mouse embryo fibroblast cell line) whole cell lysate with ab214423 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab214423 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1,000 dilution

    Lane 1: NIH/3T3 (mouse embryonic fibroblast) grown in serum-free media overnight, then treated with 50 ng/ml BMP2 for 30 minutes, whole cell lysate 10 μg (Input). 
    Lane 2: ab214423 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 ab214423 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: 5% NFDM/TBST.

References

This product has been referenced in:

  • Pei DD  et al. Contribution of Mitophagy to Cell-Mediated Mineralization: Revisiting a 50-Year-Old Conundrum. Adv Sci (Weinh) 5:1800873 (2018). Read more (PubMed: 30356983) »
See 1 Publication for this product

Customer reviews and Q&As

Application
Western blot
Sample
Mouse Tissue lysate - whole (lung, liver)
Gel Running Conditions
Reduced Denaturing (12)
Loading amount
40 µg
Specification
lung, liver
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Jul 08 2019

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
For licensing inquiries, please contact partnerships@abcam.com

Sign up