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PhosphoTracer IKKα pSer176/180 + Total ELISA Kit (ab119662)

Abreviews (1)Q&A (2)

Overview

  • Product namePhosphoTracer IKKα pSer176/180 + Total ELISA Kit
  • Tests
    1 x 96 well plate
  • Sample type
    Cell culture extracts
  • Assay typeSandwich
  • Assay time
    1h 15m
  • Species reactivity
    Reacts with: Mouse, Human
    Predicted to work with: Rat
  • Product overview

    PhosphoTracer assays use a traditional immuno-sandwich format, but with a major difference both the analyte and the assay reagents are added to the PhosphoTracer assay microplate at the same time. After a short incubation period, unbound assay reagents and analytes are washed away, and immuno-complexes containing both antibodies are detected. The process can take as little as 60 minutes to complete.

    PhosphoTracer kits also allow a higher degree of assay flexibility. In contrast to other ELISA formats, no antibodies are present on the assay microplate itself, so assays for several different targets can be performed in different wells on the same microplate. Simply mix the lysate with your target reagents of choice, using the microplate configuration of your choice.

    A whole new way of performing cellular assays, PhosphoTracer takes the hard work out of running a standard ELISA, while still giving the high quality results expected from a sandwich immunoassay. Fully self-contained kits are supplied in convenient 96-well packs. Simple to use and highly sensitive PhosphoTracer kits are designed to get results, fast.

     

    Abcam's PhosphoTracer IKKalpha assay kits detect endogenous levels of IKKalpha (GenBank Accession NP_001269) in cellular lysates. Phospho-IKKalpha assay kits detect IKKalpha only when phosphorylated at Ser176/180. Total IKKalpha assay kits detect IKKalpha irrespective of phosphorylation status.



    The substrate used with the HRP conjugated detection antibody is a combination of 10-Acetyl-3,7-dihydroxyphenoxazine (ADHP) (wavelength exc/em = 530-540nm / 590-600nm), a highly sensitive and stable substrate for HRP) and ADHP Dilution Buffer (a stabilized H2O2 solution). Learn more about the fluorogenic substrate, ADHP.

  • Notes

    Sensitivity: Phospho-IKKalpha: 0.25ng/ml (tested with rhIKKalpha), IKKalpha: 0.5ng/ml (tested with rhIKKalpha).

    Range: Phospho-IKKalpha: 0.25 - 250 ng/ml (tested with rhIKKalpha), IKKalpha: 0.5 - 250 ng/ml (tested with rhIKKalpha).

  • Tested applicationsSandwich ELISA more details

Properties

  • FunctionActs as part of the IKK complex in the conventional pathway of NF-kappa-B activation and phosphorylates inhibitors of NF-kappa-B thus leading to the dissociation of the inhibitor/NF-kappa-B complex and ultimately the degradation of the inhibitor. As part of the non-canonical pathway of NF-kappa-B activation, the MAP3K14-activated CHUK/IKKA homodimer phosphorylates NFKB2/p100 associated with RelB, inducing its proteolytic processing to NFKB2/p52 and the formation of NF-kappa-B RelB-p52 complexes. Also phosphorylates NCOA3. Phosphorylates 'Ser-10' of histone H3 at NF-kappa-B-regulated promoters during inflammatory responses triggered by cytokines.
  • Tissue specificityWidely expressed.
  • Involvement in diseaseDefects in CHUK are the cause of cocoon syndrome (COCOS) [MIM:613630]; also known as fetal encasement syndrome. COCOS is a lethal syndrome characterized by multiple fetal malformations including defective face and seemingly absent limbs, which are bound to the trunk and encased under the skin.
  • Sequence similaritiesBelongs to the protein kinase superfamily. Ser/Thr protein kinase family. I-kappa-B kinase subfamily.
    Contains 1 protein kinase domain.
  • Post-translational
    modifications    Phosphorylated by MAP3K14/NIK, AKT and to a lesser extent by MEKK1, and dephosphorylated by PP2A. Autophosphorylated.
    Acetylation of Thr-179 by Yersinia yopJ prevents phosphorylation and activation, thus blocking the I-kappa-B signaling pathway.
  • Cellular localizationCytoplasm. Nucleus. Shuttles between the cytoplasm and the nucleus.

    • Target information above from: UniProt accession O15111 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
    • Alternative names
        chukConserved helix-loop-helix ubiquitous kinaseI-kappa-B kinase 1
        I-kappa-B kinase alphaIkappaB kinaseIkB kinase alpha subunitIkBKAIKK a kinaseIKK-AIKK-alphaIKK1IKKAIKKA_HUMANInhibitor of nuclear factor kappa-B kinase subunit alphaNFKBIKANuclear factor NF-kappa-B inhibitor kinase alphaNuclear factor NFkappaB inhibitor kinase alphaTCF-16TCF16Transcription factor 16
      see all
  • Database links

      • Applications

      Our Abpromise guarantee covers the use of ab119662 in the following tested applications.

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

      Application Notes
      Sandwich ELISA sELISA. Tested in HeLa.

      PhosphoTracer IKKα pSer176/180 + Total ELISA Kit images

      • Sandwich ELISA - PhosphoTracer IKKalpha (pS176/180) + total IKKa ELISA kit (ab119662)
        Sandwich ELISA - PhosphoTracer IKKalpha (pS176/180) + total IKKa ELISA kit (ab119662)
        Using the IKKalpha assay kits, a significant stimulation of IKKalpha phosphorylation at Ser176/180 is detected in HeLa cells treated with TNF alpha for 5 minutes compared with untreated cells, while no change in total IKKalpha levels is observed over this time period.
      • Sandwich ELISA - PhosphoTracer IKKalpha (pS176/180) + total IKKa ELISA kit (ab119662)
        Sandwich ELISA - PhosphoTracer IKKalpha (pS176/180) + total IKKa ELISA kit (ab119662)
        A dilution series of recombinant active IKKalpha was prepared by diluting to various concentrations in Lysis Mix, containing 0.1% BSA. These dilution series were analyzed using the phospho-IKKalpha assay. Limit of detection of active rhIKKalpha was around 250 pg/mL. No cross-reactivity with active rhIKKB was detected over this concentration range.

      Protocols

      References for PhosphoTracer IKKα pSer176/180 + Total ELISA Kit (ab119662)

      ab119662 has not yet been referenced specifically in any publications.

      Publishing research using ab119662 ? Please let us know so that we can cite the reference in this datasheet

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      Abreviews

      Sample: Human Cell (multiple myeloma)
      Specification: multiple myeloma
      Amount of Sample: 1 µg
      Type of Kit Used: ELISA
      Positive control: supplied in kit
      Did you use the Abcam Kit protocol?: Yes

      Additional data
      Additional Notes: Kit worked well with the positive control supplied, but was not sensitive enough to detect phospho-IKKa in my samples.
      Abcam response: Thank you for submitting your Abreview. This ELISA kit is able to detect Phospho-IKKalpha ranging from 0.25 - 250 ng/ml and IKKalpha ranging from 0.5 - 250 ng/ml. In our protocol, we recommend adding 50 uL of cell lysate at a concentration of 0.1 - 0.5 mg/mL. This would correspond to at least 5 ug of cell lysate per well. You should be able to detect the IKKalpha in your samples if you load more lysate per well.
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      Submitted Feb 14 2012

      Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"