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    rb-phospho-s249-antibody-ab4788.pdf

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Cell Biology Cell Cycle Cell Cycle Inhibitors Rb
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Anti-Rb (phospho S249) antibody (ab4788)

  • Datasheet
Reviews (1)Q&A (1)References (2)

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Western blot - Anti-Rb (phospho S249) antibody (ab4788)

    Key features and details

    • Rabbit polyclonal to Rb (phospho S249)
    • Suitable for: WB
    • Reacts with: Human
    • Isotype: IgG

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    Recombinant Human Rb protein (ab83205)

    View more associated products

    Overview

    • Product name

      Anti-Rb (phospho S249) antibody
      See all Rb primary antibodies
    • Description

      Rabbit polyclonal to Rb (phospho S249)
    • Host species

      Rabbit
    • Tested Applications & Species

      Application Species
      WB
      Human
      See all applications and species data
    • Immunogen

      Synthetic peptide (Human) derived from a region of human Rb that contains serine 249 and threonine 252 (based on Swiss Protein database, accession number P06400). The sequence is conserved in human, mouse and rat.

    • Positive control

      • Jurkat cells in high growth phase.

    Properties

    • Form

      Liquid
    • Storage instructions

      Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
    • Storage buffer

      pH: 7.30
      Preservative: 0.05% Sodium azide
      Constituents: PBS, 50% Glycerol (glycerin, glycerine), 0.1% BSA
    • Concentration information loading...
    • Purity

      Immunogen affinity purified
    • Purification notes

      Purified from rabbit serum by sequential epitope-specific chromatography. The antibody has been negatively preadsorbed using a non-phosphopeptide corresponding to the site of phosphorylation to remove antibody that is reactive with non-phosphorylated Rb protein. The final product is generated by affinity chromatography using a Rb-derived peptide that is phosphorylated at serine 249 and threonine 252.
    • Clonality

      Polyclonal
    • Isotype

      IgG
    • Research areas

      • Cell Biology
      • Cell Cycle
      • Cell Cycle Inhibitors
      • Rb
      • Epigenetics and Nuclear Signaling
      • Transcription
      • Cancer susceptibility
      • Tumor Suppressors
      • Cancer
      • Oncoproteins/suppressors
      • Tumor suppressors
      • Rb family

    Associated products

    • Compatible Secondaries

      • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
      • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
    • Isotype control

      • Rabbit IgG, polyclonal - Isotype Control (ChIP Grade) (ab171870)
    • Positive Controls

      • Jurkat whole cell lysate (ab7899)
    • Recombinant Protein

      • Recombinant Human Rb protein (ab83205)

    Applications

    The Abpromise guarantee

    Our Abpromise guarantee covers the use of ab4788 in the following tested applications.

    The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

    Guaranteed

    Tested applications are guaranteed to work and covered by our Abpromise guarantee.

    Predicted

    Predicted to work for this combination of applications and species but not guaranteed.

    Incompatible

    Does not work for this combination of applications and species.

    Application Species
    WB
    Human
    All applications
    Mouse
    Rat
    Chicken
    Application Abreviews Notes
    WB
    Notes
  • Application notes
    WB: 1/1000. Detects a band of approximately 120 kDa (predicted molecular weight: 106 kDa).
    Can be blocked with Rb peptide - phospho S249 + T252 (phospho and non-phospho pair).

    Not tested in other applications.
    Optimal dilutions/concentrations should be determined by the end user.
  • Target

    • Function

      Key regulator of entry into cell division that acts as a tumor suppressor. Promotes G0-G1 transition when phosphorylated by CDK3/cyclin-C. Acts as a transcription repressor of E2F1 target genes. The underphosphorylated, active form of RB1 interacts with E2F1 and represses its transcription activity, leading to cell cycle arrest. Directly involved in heterochromatin formation by maintaining overall chromatin structure and, in particular, that of constitutive heterochromatin by stabilizing histone methylation. Recruits and targets histone methyltransferases SUV39H1, KMT5B and KMT5C, leading to epigenetic transcriptional repression. Controls histone H4 'Lys-20' trimethylation. Inhibits the intrinsic kinase activity of TAF1. Mediates transcriptional repression by SMARCA4/BRG1 by recruiting a histone deacetylase (HDAC) complex to the c-FOS promoter. In resting neurons, transcription of the c-FOS promoter is inhibited by BRG1-dependent recruitment of a phospho-RB1-HDAC1 repressor complex. Upon calcium influx, RB1 is dephosphorylated by calcineurin, which leads to release of the repressor complex (By similarity). In case of viral infections, interactions with SV40 large T antigen, HPV E7 protein or adenovirus E1A protein induce the disassembly of RB1-E2F1 complex thereby disrupting RB1's activity.
    • Tissue specificity

      Expressed in the retina.
    • Involvement in disease

      Childhood cancer retinoblastoma
      Bladder cancer
      Osteogenic sarcoma
    • Sequence similarities

      Belongs to the retinoblastoma protein (RB) family.
    • Domain

      The Pocket domain binds to the threonine-phosphorylated domain C, thereby preventing interaction with heterodimeric E2F/DP transcription factor complexes.
    • Post-translational
      modifications

      Phosphorylated by CDK6 and CDK4, and subsequently by CDK2 at Ser-567 in G1, thereby releasing E2F1 which is then able to activate cell growth. Dephosphorylated at the late M phase. SV40 large T antigen, HPV E7 and adenovirus E1A bind to the underphosphorylated, active form of pRb. Phosphorylation at Thr-821 and Thr-826 promotes interaction between the C-terminal domain C and the Pocket domain, and thereby inhibits interactions with heterodimeric E2F/DP transcription factor complexes. Dephosphorylated at Ser-795 by calcineruin upon calcium stimulation. CDK3/cyclin-C-mediated phosphorylation at Ser-807 and Ser-811 is required for G0-G1 transition. Phosphorylated by CDK1 and CDK2 upon TGFB1-mediated apoptosis.
      N-terminus is methylated by METTL11A/NTM1 (By similarity). Monomethylation at Lys-810 by SMYD2 enhances phosphorylation at Ser-807 and Ser-811, and promotes cell cycle progression. Monomethylation at Lys-860 by SMYD2 promotes interaction with L3MBTL1.
      Acetylation at Lys-873 and Lys-874 regulates subcellular localization, at least during keratinocytes differentiation.
    • Cellular localization

      Nucleus.
    • Target information above from: UniProt accession P06400 The UniProt Consortium
      The Universal Protein Resource (UniProt) in 2010
      Nucleic Acids Res. 38:D142-D148 (2010) .

      Information by UniProt
    • Database links

      • Entrez Gene: 386582 Chicken
      • Entrez Gene: 5925 Human
      • Entrez Gene: 19645 Mouse
      • Entrez Gene: 24708 Rat
      • Omim: 614041 Human
      • SwissProt: Q90600 Chicken
      • SwissProt: P06400 Human
      • SwissProt: P13405 Mouse
      • SwissProt: P33568 Rat
      • Unigene: 408528 Human
      • Unigene: 273862 Mouse
      • Unigene: 55115 Rat
      see all
    • Alternative names

      • Exon 17 tumor GOS561 substitution mutation causes premature stop antibody
      • GOS563 exon 17 substitution mutation causes premature stop antibody
      • OSRC antibody
      • Osteosarcoma antibody
      • p105-Rb antibody
      • P105RB antibody
      • PP105 antibody
      • pp110 antibody
      • PPP1R130 antibody
      • pRb antibody
      • Prepro retinoblastoma associated protein antibody
      • Protein phosphatase 1 regulatory subunit 130 antibody
      • Rb antibody
      • RB transcriptional corepressor 1 antibody
      • RB_HUMAN antibody
      • RB1 antibody
      • RB1 gene antibody
      • Retinoblastoma 1 antibody
      • Retinoblastoma suspectibility protein antibody
      • Retinoblastoma-associated protein antibody
      see all

    Images

    • Western blot - Anti-Rb (phospho S249) antibody (ab4788)
      Western blot - Anti-Rb (phospho S249) antibody (ab4788)
      Peptide Competition: Cell extracts prepared from Jurkat cells in high growth phase were resolved by SDS-PAGE on a 10% Tris-glycine gel and transferred to PVDF. Membranes were incubated with 0.50 µg/mL ab4788, following prior incubation in the absence of the phosphopeptide immunogen (1), a generic phosphoserine containing peptide (2), a generic phosphothreonine containing peptide (3), the non-phosphopeptide corresponding to the phosphopeptide immunogen (4), or the presence of the phosphopeptide immunogen (5). After washing, membranes were incubated with goat F(ab’)2 anti-rabbit IgG alkaline phosphatase and bands were detected using the Tropix WesternStar detection method. The data show that only the phosphopeptide corresponding to this site blocks the antibody signal, therefore demonstrating the specificity of the ab4788 antibody for this epitope. Peptide Competition: Cell extracts prepared from Jurkat cells in high growth phase were resolved by SDS-PAGE on a 1

    Protocols

    • Peptide Competition Experiment
    • Western blotting protocol

    Click here to view the general protocols

    Datasheets and documents

    • Datasheet
  • References (2)

    Publishing research using ab4788? Please let us know so that we can cite the reference in this datasheet.

    ab4788 has been referenced in 2 publications.

    • Pérez-Morales J  et al. Hyper-phosphorylation of Rb S249 together with CDK5R2/p39 overexpression are associated with impaired cell adhesion and epithelial-to-mesenchymal transition: Implications as a potential lung cancer grading and staging biomarker. PLoS One 13:e0207483 (2018). PubMed: 30452490
    • Lee YC  et al. Macrophage inflammatory protein-3alpha influences growth of K562 leukemia cells in co-culture with anticancer drug-pretreated HS-5 stromal cells. Toxicology 249:116-22 (2008). WB ; Human . PubMed: 18538911

    Customer reviews and Q&As

    Show All Reviews Q&A
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    1-2 of 2 Abreviews or Q&A

    Immunocytochemistry/ Immunofluorescence abreview for Anti-Rb (phospho S249) antibody

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (HeLa)
    Permeabilization
    Yes - 0.5% Triton X-100
    Specification
    HeLa
    Fixative
    Paraformaldehyde
    Read More

    Dr. Kirk Mcmanus

    Verified customer

    Submitted Jul 03 2018

    Question

    I have some problem with ab4788 (anti retinoblastoma) you send me two weeks ago. First of all it was supposed to be 50ul but it was only 20ul. Second, it doesn't detect any band even in the positive control (20ug of jurkat lysate, ab7899). I tried two times with two different incubations (2 hours at room temperature as you suggest in your protocol) and overnight at 4 degrees with no results. I hope you could resolve these problems.

    Read More

    Abcam community

    Verified customer

    Asked on Feb 03 2005

    Answer

    Thank you for your email and I'm sorry to hear about the difficulty you are experiencing with ab4788. I apologize for the shortage that you received and can certainly send you a replacement vial free of charge. Can you please tell me the batch number that you received (it is located on the vial)? Also, what was the Abcam order number or purchase number that was used? Thank you, and I look forward to hearing from you.

    Read More

    Abcam Scientific Support

    Answered on Feb 04 2005

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