Overview

  • Product name

    Human Antithrombin III / ATIII ELISA Kit
    See all Antithrombin III/ATIII kits
  • Detection method

    Colorimetric
  • Precision

    Intra-assay
    Sample n Mean SD CV%
    Serum 8 2.9%
    Inter-assay
    Sample n Mean SD CV%
    Serum 3 7.9%
  • Sample type

    Cell culture supernatant, Urine, Serum, Hep Plasma, EDTA Plasma, Cit plasma
  • Assay type

    Sandwich (quantitative)
  • Sensitivity

    0.33 ng/ml
  • Range

    3.13 ng/ml - 200 ng/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Cell culture supernatant 120 106% - 133%
    Milk 113 100% - 126%
    Urine 87 80% - 93%
    Serum 91 79% - 106%
    Cell culture media 110 97% - 122%
    Hep Plasma 106 97% - 121%
    EDTA Plasma 100 102% - 110%
    Cit plasma 107 79% - 106%

  • Assay time

    1h 30m
  • Assay duration

    One step assay
  • Species reactivity

    Reacts with: Human
  • Product overview

    Antithrombin III in vitro SimpleStep ELISA® (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of human Antithrombin III protein in serum, plasma, urine, milk, and cell culture supernatant 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.


    Antithrombin-III (ATIII), also known as Serpin C1, is a 58 kDa extracellular glycoprotein belonging to the Serpin serine protease inhibitor superfamily. Antithrombin III functions primarily as a blood anticoagulant but also modulates inflammatory cytokine expression. Human Antithrombin-III is synthesized in the liver and is secreted as a 432 amino acid (aa) mature protein. Most circulating Antithrombin-III (90-95%) is the fully glycosylated α-isoform while the partially glycosylated β-isoform constitutes 5-10%. The biological activity of Antithrombin-III is inherently low and is enhanced up to 1000-fold in the presence of heparin or heparin-like glycosaminoglycans. The β-isoform of Antithrombin-III has a higher affinity for heparin. Mature human Antithrombin-III shares 90% sequence homology with mature mouse Antithrombin-III. Human Antithrombin-III inactivates thrombin and other blood coagulation factors. Human adult Antithrombin-III levels in plasma are relatively constant. However, reduction of Antithrombin-III is observed in individuals with diabetes or sepsis. Neonates also exhibit low levels of Serpin C1 in plasma.

  • Tested applications

    Suitable for: Sandwich ELISAmore details
  • 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
    50X Cell Extraction Enhancer Solution (ab193971) 1 x 1ml
    5X Cell Extraction Buffer PTR (ab193970) 1 x 10ml
    Antibody Diluent 5BC 1 x 6ml
    Denaturant 1 x 500µl
    10X Human Antithrombin III Capture Antibody 1 x 600µl
    10X Human Antithrombin III Detector Antibody 1 x 600µl
    Human Antithrombin III Lyophilized Recombinant Protein 1 x 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

    Most important serine protease inhibitor in plasma that regulates the blood coagulation cascade. AT-III inhibits thrombin as well as factors IXa, Xa and XIa. Its inhibitory activity is greatly enhanced in the presence of heparin.
  • Tissue specificity

    Found in plasma.
  • Involvement in disease

    Defects in SERPINC1 are the cause of antithrombin III deficiency (AT3D) [MIM:613118]. AT3D is an important risk factor for hereditary thrombophilia, a hemostatic disorder characterized by a tendency to recurrent thrombosis. AT3D is classified into 4 types. Type I: characterized by a 50% decrease in antigenic and functional levels. Type II: has defects affecting the thrombin-binding domain. Type III: alteration of the heparin-binding domain. Plasma AT-III antigen levels are normal in type II and III. Type IV: consists of miscellaneous group of unclassifiable mutations.
  • Sequence similarities

    Belongs to the serpin family.
  • Post-translational
    modifications

    Phosphorylation sites are present in the extracelllular medium.
  • Cellular localization

    Secreted > extracellular space.
  • Information by UniProt
  • Alternative names

    • ANT3_HUMAN
    • Antithrombin
    • Antithrombin III
    • Antithrombin-III
    • AntithrombinIII
    • AT 3
    • AT III
    • AT3
    • AT3D
    • ATIII
    • Heparin cofactor I
    • MGC22579
    • Serine (or cysteine) proteinase inhibitor clade C (antithrombin) member 1
    • Serine cysteine proteinase inhibitor clade C member 1
    • Serine proteinase inhibitor clade C member 1
    • Serpin C1
    • Serpin family C member 1
    • Serpin peptidase inhibitor clade C (antithrombin) member 1
    • SERPINC1
    • THPH7
    see all
  • Database links

Applications

Our Abpromise guarantee covers the use of ab222507 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
Sandwich ELISA Use at an assay dependent concentration.

Images

  • 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.

     

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

  • The concentrations of Antithrombin III were measured in duplicates, interpolated from the Antithrombin III standard curves and corrected for sample dilution. Undiluted samples are as follows: serum 1:4,000 plasma (citrate) 1:8,000, plasma (EDTA) 1:4,000, and plasma (heparin) 1:4,000. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Antithrombin III concentration was determined to be 157.69 µg/mL in neat serum, 279.95 µg/mL in neat plasma (citrate), 160.27 µg/mL in neat plasma (EDTA), and 224.88 µg/mL in neat plasma (heparin).

  • The concentrations of Antithrombin III were measured in duplicates, interpolated from the Antithrombin III standard curves and corrected for sample dilution. Undiluted samples are as follows: milk 3.13%, Hep G2 supernatant 6.25%, and urine 6.25%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Antithrombin III concentration was determined to be 810.16 ng/mL in neat milk, 218.86 ng/mL in neat Hep G2 supernatant, and 618.60 ng/mL in neat spiked urine.

  • Interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Antithrombin III concentration was determined to be 77.20 µg/mL with a range of 34.09 – 121.12 µg/mL.

Protocols

References

ab222507 has not yet been referenced specifically in any publications.

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