SimpleStep

Human Thrombomodulin ELISA Kit (ab214029)

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Overview

  • Product name

    Human Thrombomodulin ELISA Kit
    See all Thrombomodulin kits

  • Detection method

    Colorimetric

  • Precision

    Intra-assay
    Sample n Mean SD CV%
    Human serum 8 3.1%
    Inter-assay
    Sample n Mean SD CV%
    Human serum 3 9.4%

  • Sample type

    Cell culture supernatant, Urine, Serum, Cell culture extracts, Tissue Extracts, Hep Plasma, EDTA Plasma, Cit plasma

  • Assay type

    Sandwich (quantitative)

  • Sensitivity

    3 pg/ml

  • Range

    20.3 pg/ml - 1300 pg/ml

  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Cell culture supernatant 90 88% - 91%
    Urine 106 102% - 112%
    Serum 97 96% - 99%
    Cell culture extracts 94 92% - 96%
    Tissue Extracts 105 101% - 110%
    Hep Plasma 99 98% - 100%
    EDTA Plasma 96 94% - 98%
    Cit plasma 96 92% - 98%

  • Assay time

    1h 30m

  • Assay duration

    One step assay

  • Species reactivity

    Reacts with: Human
    Does not react with: Cow

  • Product overview

    Thrombomodulin in vitro SimpleStep ELISA® (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of Thrombomodulin protein in serum, plasmas, urine, cell culture supernatants, and cell and tissue extract samples.


    The SimpleStep ELISA® employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material. TMB substrate is added and during incubation is catalyzed by HRP, generating blue coloration. This reaction is then stopped by addition of Stop Solution completing any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.


    Sensitivity:
    Samples in Sample Diluent NS: 11 pg/mL
    Samples in 1X Cell Extraction Buffer PTR: 3 pg/mL

  • Notes

    Human Thrombomodulin is a 557 amino acid type I transmembrane protein, consisting of six epidermal growth factor (EGF)-like domains and is highly glycosylated. Thrombomodulin is an important component in the anti-coagulation system by functioning as a cell surface receptor for thrombin, and has also been shown to play a role in anti-inflammation, cell adhesion, and proliferation. The Thrombomodulin - thrombin complex activates protein C and thrombin-activatable fibrinolysis inhibitor (TAFI); protein C degrades coagulation factors Va and VIIIa, which regulates thrombin resulting in inhibition of coagulation. The extracellular portion of Thrombomodulin has been shown to be released from the cell surface due to protein cleavage during vascular endothelial cell injury.

  • Platform

    Pre-coated microplate (12 x 8 well strips)

Properties

  • Storage instructions

    Store at +4°C. Please refer to protocols.
  • Components 1 x 96 tests
    10X Wash Buffer PT (ab206977) 1 x 20ml
    20X Human Thrombomodulin Detector Antibody 1 x 300µl
    50X Cell Extraction Enhancer Solution (ab193971) 1 x 1ml
    5X Cell Extraction Buffer PTR (ab193970) 1 x 10ml
    Antibody Diluent 4BI 1 x 6ml
    Human Thrombomodulin Capture Antibody (Lyophilized) 1 vial
    Human Thrombomodulin Lyophilized Recombinant Protein 2 vials
    Plate Seals 1 unit
    Sample Diluent NS (ab193972) 1 x 50ml
    SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit
    Stop Solution 1 x 12ml
    TMB Development Solution 1 x 12ml

  • Research areas

  • Function

    Thrombomodulin is a specific endothelial cell receptor that forms a 1:1 stoichiometric complex with thrombin. This complex is responsible for the conversion of protein C to the activated protein C (protein Ca). Once evolved, protein Ca scissions the activated cofactors of the coagulation mechanism, factor Va and factor VIIIa, and thereby reduces the amount of thrombin generated.

  • Tissue specificity

    Endothelial cells are unique in synthesizing thrombomodulin.

  • Involvement in disease

    Defects in THBD are the cause of thrombophilia due to thrombomodulin defect (THR-THBD) [MIM:188040]. A hemostatic disorder characterized by a tendency to thrombosis.
    Defects in THBD are a cause of susceptibility to hemolytic uremic syndrome atypical type 6 (AHUS6) [MIM:612926]. An atypical form of hemolytic uremic syndrome. It is a complex genetic disease characterized by microangiopathic hemolytic anemia, thrombocytopenia, renal failure and absence of episodes of enterocolitis and diarrhea. In contrast to typical hemolytic uremic syndrome, atypical forms have a poorer prognosis, with higher death rates and frequent progression to end-stage renal disease. Note=Susceptibility to the development of atypical hemolytic uremic syndrome can be conferred by mutations in various components of or regulatory factors in the complement cascade system. Other genes may play a role in modifying the phenotype.

  • Sequence similarities

    Contains 1 C-type lectin domain.
    Contains 6 EGF-like domains.

  • Post-translational
    modifications

    N-glycosylated.
    The iron and 2-oxoglutarate dependent 3-hydroxylation of aspartate and asparagine is (R) stereospecific within EGF domains.

  • Cellular localization

    Membrane.
  • Information by UniProt

  • Alternative names

    • AHUS 6
    • AHUS6
    • BDCA 3
    • BDCA3
    • CD 141
    • CD141
    • CD141 antigen
    • Fetomodulin
    • Thbd
    • THPH12
    • THRM
    • Thrombomodulin
    • TM
    • TRBM_HUMAN
    see all

  • Database links

    Images

    • Other - Human Thrombomodulin ELISA Kit (ab214029)
      Other - Human Thrombomodulin ELISA Kit (ab214029)

      SimpleStep ELISA technology allows the formation of the antibody-antigen complex in one single step, reducing assay time to 90 minutes. Add samples or standards and antibody mix to wells all at once, incubate, wash, and add your final substrate. See protocol for a detailed step-by-step guide.

       

    • Human Thrombomodulin standard curve comparison data.
      Human Thrombomodulin standard curve comparison data.

      Standard curve comparison between human Thrombomodulin SimpleStep ELISA® kit and traditional ELISA kit from leading competitor. SimpleStep ELISA kit shows a 8-fold increase in sensitivity.

    • Example of Human Thrombomodulin standard curve in Sample Diluent NS.
      Example of Human Thrombomodulin standard curve in Sample Diluent NS.

      Background-subtracted data values (mean +/- SD) are graphed.

    • Example of Human Thrombomodulin standard curve in 1X Cell Extraction Buffer PTR.
      Example of Human Thrombomodulin standard curve in 1X Cell Extraction Buffer PTR.

      Background-subtracted data values (mean +/- SD) are graphed.

    • Interpolated concentrations of native Thrombomodulin in human serum, and plasma samples.
      Interpolated concentrations of native Thrombomodulin in human serum, and plasma samples.

      The concentrations of Thrombomodulin were measured in duplicate, interpolated from the Thrombomodulin standard curve and corrected for sample dilution. Undiluted samples are as follows: serum 25%, plasma (citrate) 25%, plasma (EDTA) 25% and plasma (heparin) 25%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Thrombomodulin concentration was determined to be 4.8 ng/mL in serum, 3.9 ng/mL in plasma (citrate), 3.8 ng/mL in plasma (EDTA) and 4.3 ng/mL in plasma (heparin).

    • Interpolated concentrations of spiked recombinant Thrombomodulin in HepG2 cell culture supernatant samples.
      Interpolated concentrations of spiked recombinant Thrombomodulin in HepG2 cell culture supernatant samples.

      The concentrations of Thrombomodulin were measured in duplicate, interpolated from the Thrombomodulin standard curve and corrected for sample dilution. Undiluted sample is at 45%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The native Thrombomodulin concentration was undetectable in the null spike control run in parallel with the spiked sample.

    • Interpolated concentrations of native Thrombomodulin in HUVEC and human liver homogenate extracts based on a 1,000 µg/mL extract load.
      Interpolated concentrations of native Thrombomodulin in HUVEC and human liver homogenate extracts based on a 1,000 µg/mL extract load.

      The concentrations of Thrombomodulin were measured in duplicate and interpolated from the Thrombomodulin standard curve and corrected for sample dilution. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Thrombomodulin concentration was determined to be 1.2 ng/mL in HUVEC extract and 8.0 ng/mL in liver homogenate extract.

    • Serum from ten individual healthy human male donors was measured in duplicate.
      Serum from ten individual healthy human male donors was measured in duplicate.

      Interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Thrombomodulin concentration was determined to be 5.1 ng/mL with a range of 3.6 – 6.8 ng/mL.

    • Serum from ten individual healthy human female donors was measured in duplicate.
      Serum from ten individual healthy human female donors was measured in duplicate.

      Interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean target concentration was determined to be 5.1 ng/mL with a range of 2.7 – 9.9 ng/mL.

    Protocols

    References

    This product has been referenced in:

    • Kamtchum-Tatuene J  et al. HIV is associated with endothelial activation despite ART, in a sub-Saharan African setting. Neurol Neuroimmunol Neuroinflamm 6:e531 (2019). ELISA . Read more (PubMed: 30697583) »
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