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  1. Link

    products/primary-antibodies/cblb-antibody-246c5a-ab54362.pdf

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Immunology Innate Immunity Macrophage / Inflamm.
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Validated using a knockout cell line

Anti-CBLB antibody [246C5a] (ab54362)

  • Datasheet
  • SDS
Reviews (1) Submit a question References (2)

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CBLB antibody [246C5a] (ab54362)
  • Western blot - Anti-CBLB antibody [246C5a] (ab54362)
  • Western blot - Anti-CBLB antibody [246C5a] (ab54362)

Key features and details

  • Mouse monoclonal [246C5a] to CBLB
  • Suitable for: IHC-P, WB
  • Knockout validated
  • Reacts with: Mouse, Human
  • Isotype: IgG2a

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Overview

  • Product name

    Anti-CBLB antibody [246C5a]
    See all CBLB primary antibodies
  • Description

    Mouse monoclonal [246C5a] to CBLB
  • Host species

    Mouse
  • Tested applications

    Suitable for: IHC-P, WBmore details
  • Species reactivity

    Reacts with: Mouse, Human
  • Immunogen

    Recombinant C terminal fragment of Human CBLB.

  • Positive control

    • WB: CTLL-2, HeLa and MCF7 cell lysates
  • General notes

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.

    If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As

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 long term.
  • Storage buffer

    pH: 7.40
    Preservative: 0.05% Sodium azide
    Constituents: 1% BSA, PBS
  • Concentration information loading...
  • Purity

    Protein G purified
  • Purification notes

    ab54362 was purified using protein G column chromatography from culture supernatant of hybridoma cultured in a medium containing bovine IgG depleted (approximately 95%) fetal bovine serum and filtered through a 0.22µm membrane.
  • Clonality

    Monoclonal
  • Clone number

    246C5a
  • Isotype

    IgG2a
  • Research areas

    • Immunology
    • Innate Immunity
    • Macrophage / Inflamm.
    • Cell Biology
    • Proteolysis / Ubiquitin
    • Proteasome / Ubiquitin
    • Ubiquitin E3 Enzymes
    • RING Finger E3 Ligase
    • Immunology
    • Immune System Diseases
    • Autoimmune
    • Cancer
    • Oncoproteins/suppressors
    • Oncoproteins
    • Signal transducers

Associated products

  • Compatible Secondaries

    • Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) (ab150113)
    • Goat Anti-Mouse IgG H&L (HRP) (ab205719)
  • Isotype control

    • Mouse IgG2a, kappa monoclonal [MG2a-53] - Isotype control (ab18415)
  • KO cell lines

    • Human CBLB knockout HeLa cell line (ab264711)
  • KO cell lysates

    • Human CBLB knockout HeLa cell lysate (ab258342)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab54362 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
IHC-P
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
WB (1)
Use at an assay dependent concentration. Predicted molecular weight: 109 kDa.
Notes
IHC-P
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
WB
Use at an assay dependent concentration. Predicted molecular weight: 109 kDa.

Target

  • Relevance

    Function: E3 ubiquitin-protein ligase which accepts ubiquitin from specific E2 ubiquitin-conjugating enzymes, and transfers it to substrates, generally promoting their degradation by the proteasome. Negatively regulates TCR (T-cell receptor), BCR (B-cell receptor) and FCER1 (high affinity immunoglobulin epsilon receptor) signal transduction pathways. In naive T-cells, inhibits VAV1 activation upon TCR engagement and imposes a requirement for CD28 costimulation for proliferation and IL-2 production. Also acts by promoting PIK3R1/p85 ubiquitination, which impairs its recruitment to the TCR and subsequent activation. In activated T-cells, inhibits PLCG1 activation and calcium mobilization upon restimulation and promotes anergy. In B-cells, acts by ubiquitinating SYK and promoting its proteasomal degradation. Slightly promotes SRC ubiquitination. May be involved in EGFR ubiquitination and internalization. May be functionally coupled with the E2 ubiquitin-protein ligase UB2D3. Tissue specificity: Expressed in placenta, heart, lung, kidney, spleen, ovary and testis, as well as fetal brain and liver and hematopoietic cell lines, but not in adult brain, liver, pancreas, salivary gland, or skeletal muscle. Present in lymphocytes (at protein level). Pathway: Protein modification; protein ubiquitination. Similarity: Contains 1 Cbl-PTB (Cbl-type phosphotyrosine-binding) domain. Contains 1 RING-type zinc finger. Contains 1 UBA domain. Domain: The N-terminus is composed of the phosphotyrosine binding (PTB) domain, a short linker region and the RING-type zinc finger. The PTB domain, which is also called TKB (tyrosine kinase binding) domain, is composed of three different subdomains: a four-helix bundle (4H), a calcium-binding EF hand and a divergent SH2 domain. The RING-type zinc finger domain mediates binding to an E2 ubiquitin-conjugating enzyme. The UBA domain interacts with poly-ubiquitinated proteins. PTM: Phosphorylated on tyrosine and serine residues upon TCR or BCR activation, and upon various types of cell stimulation. Auto-ubiquitinated upon EGF-mediated cell activation or upon T-cell costimulation by CD28; which promotes proteasomal degradation.
  • Cellular localization

    Cytoplasmic and Nuclear
  • Database links

    • Entrez Gene: 868 Human
    • Entrez Gene: 208650 Mouse
    • Omim: 604491 Human
    • SwissProt: Q13191 Human
    • SwissProt: Q3TTA7 Mouse
    • Unigene: 430589 Human
    • Unigene: 328206 Mouse
    • Alternative names

      • AI429560 antibody
      • AI851073 antibody
      • Cas Br M (murine) ecotropic retroviral transforming sequence b antibody
      • Casitas B lineage lymphoma b antibody
      • Casitas B lineage lymphoma proto oncogene b antibody
      • Casitas B-lineage lymphoma proto-oncogene b antibody
      • Cbl b antibody
      • CBLB antibody
      • CBLB_HUMAN antibody
      • DKFZp686J10223 antibody
      • DKFZp779A0729 antibody
      • DKFZp779F1443 antibody
      • E3 ubiquitin protein ligase CBL B antibody
      • E3 ubiquitin-protein ligase CBL-B antibody
      • EC 6.3.2.- antibody
      • FLJ36865 antibody
      • FLJ41152 antibody
      • Nbla00127 antibody
      • RING finger protein 56 antibody
      • RNF56 antibody
      • SH3 binding protein CBL B antibody
      • SH3-binding protein CBL-B antibody
      • Signal transduction protein CBL B antibody
      • Signal transduction protein CBL-B antibody
      see all

    Images

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CBLB antibody [246C5a] (ab54362)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CBLB antibody [246C5a] (ab54362)

      IHC image of ab54362 staining in human T-cell lymphoma 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 ab54362, 5µg/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.

    • Western blot - Anti-CBLB antibody [246C5a] (ab54362)
      Western blot - Anti-CBLB antibody [246C5a] (ab54362)
      All lanes : Anti-CBLB antibody [246C5a] (ab54362) at 1/500 dilution

      Lane 1 : Wild-type HeLa cell lysate
      Lane 2 : CBLB knockout HeLa cell lysate
      Lane 3 : CTLL-2 cell lysate
      Lane 4 : MCF7 cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 109 kDa
      Observed band size: 130 kDa why is the actual band size different from the predicted?



      Lanes 1-4: Merged signal (red and green). Green - ab54362 observed at 130 kDa. Red - loading control, ab181602 observed at 37 kDa.

      ab54362 Anti-CBLB antibody [246C5a] was shown to specifically react with CBLB in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264711 (knockout cell lysate ab258342) was used. Wild-type and CBLB knockout samples were subjected to SDS-PAGE. ab54362 and Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) were incubated overnight at 4°C at 1 in 500 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.

       

    • Western blot - Anti-CBLB antibody [246C5a] (ab54362)
      Western blot - Anti-CBLB antibody [246C5a] (ab54362)
      Anti-CBLB antibody [246C5a] (ab54362) + immunizing recombinant protein

      Predicted band size: 109 kDa
      Observed band size: 37 kDa why is the actual band size different from the predicted?



      The molecular weight of the band on the Western blot does not correspond to the molecular weight of the natural protein as the immunizing recombinant protein fragment was used as the test antigen.

    Protocols

    • Western blot protocols
    • Immunohistochemistry protocols

    Click here to view the general protocols

    Datasheets and documents

    • SDS download

    • Datasheet download

      Download

    References (2)

    Publishing research using ab54362? Please let us know so that we can cite the reference in this datasheet.

    ab54362 has been referenced in 2 publications.

    • Singh TP  et al. Cbl-b deficiency provides protection against UVB-induced skin damage by modulating inflammatory gene signature. Cell Death Dis 9:835 (2018). PubMed: 30082827
    • Wallner S  et al. The role of the e3 ligase cbl-B in murine dendritic cells. PLoS One 8:e65178 (2013). WB ; Mouse . PubMed: 23762309

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    Western blot abreview for Anti-CBLB antibody [246C5a]

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Mouse Cell lysate - whole cell (mouse embryonic stem cells)
    Gel Running Conditions
    Reduced Denaturing (10%)
    Loading amount
    1e+007 cells
    Specification
    mouse embryonic stem cells
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°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.

    MISS. Fabiana Luise

    Verified customer

    Submitted Nov 17 2015

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