Sclerostin ELISA Assay Kit

$695.00

The Sclerostin ELISA Assay Kit is an enzyme immunoassay for the quantitative determination of sclerostin in serum or heparin plasma. The Eagle Biosciences Sclerostin ELISA Assay Kit is for research use only and not to be used in diagnostic procedures.

SKU: BI-20492 Categories: , ,

Sclerostin ELISA Assay Kit

Sclerostin ELISA Assay Kit Developed and Manufactured in Austria by Biomedica

Size: 1×96 wells
Sensitivity: LOD: (0 pmol/l + 3 SD): 3.2 pmol/l; LLOQ: <7.5 pmol/l
Dynamic Range: 0 to 240 pmol/l
Incubation Time: > 24 hours
Sample Type: Serum, Plasma (EDTA, Heparin), Urine protocol available
Sample Size: 20 µL
Alternative Names: SOST,CDD, VBCH, DAND6, SOST1
For Research Use Only
Controls Included

Product Developed and Manufactured in Austria by Biomedica

Unit conversion: 1 pg/ml = 0.044 pmol/l (MW: 22.5 kD)

Assay Principle for the Sclerostin Assay

The Sclerostin ELISA Assay Kit is a sandwich enzyme immunoassay for the quantitative determination of Sclerostin (SOST) in human serum, plasma, urine and cell culture supernatnat samples. In a first step, assay buffer is pipetted into the wells of the microtiter strips. Thereafter, standard/control/sample and detection antibody (biotinylated monoclonal mouse anti-human Sclerostin) are pipetted into the wells, which are pre-coated with polyclonal goat anti-human Sclerostin antibody. Any Sclerostin present in the standard/control/sample binds to the pre-coated antibody in the well and forms a sandwich with the detection antibody. In a washing step, all non-specific unbound material is removed. In a next step, the conjugate (strepdavidin-HRP) is pipetted into the wells and reacts with the detection antibody. After another washing step, the substrate (tetramethylbenzidine, TMB) is pipetted into the wells. The enzyme-catalyzed color reaction of the substrate is directly proportional to the amount of Sclerostin present in the sample. This color change is detectable with a standard microtiter plate reader. A dose response curve of the absorbance (optical density, OD, at 450 nm) versus standard concentration is generated, using the values obtained from the standards. The concentration of human Sclerostin in the sample is determined directly from the dose response curve. The human Sclerostin (SOST) immunoassay is an overnight, 96-well sandwich ELISA for the quantitative determination of human Sclerostin in serum, plasma, urine and cell culture supernatant. The assay employs human serum-based standards to ensure the measurement of biologically reliable data. The Sclerostin (SOST) ELISA kit uses highly purified, epitope mapped antibodies.

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Additional Information

Assay Background

Canonical Wnt-signalling plays an important role in the regulation of bone homeostasis by promoting the development of osteoblasts. Negative regulators of the Wnt pathway are important new therapeutic targets for the treatment of diseases with enhanced bone resorption. One of these molecules is Sclerostin, a 22.5 kD secreted glycoprotein, which acts by binding to the Wnt-coreceptor LRP5 thus preventing the binding of Wnt molecules. Sclerostin is nearly exclusively produced in osteocytes. Therefore it is considered a clinical marker which provides highest bone specificity.

Assay Procedure

  1. Add 150 µl ASYBUF (red cap) into each well.
  2. Add 20 µl STD/SAMPLE/CTRL (Standard/Sample/Control) in duplicate into respective well.
  3. Add 50 µl AB (biotinylated anti Sclerostin antibody, green cap, green dye) into each well, swirl gently.
  4. Cover tightly and incubate overnight (18-24 h) at room temperature (18-24°C) in the dark. Attention: Incubation higher than room temperature reduces the top-OD.
  5. Aspirate and wash wells 5x with 300 µl diluted WASHBUF (Wash buffer). After final wash, remove remaining WASHBUF by strongly tapping plate against paper towel.
  6. Add 200 µl CONJ (Conjugate, amber cap) into each well.
  7. Cover tightly and incubate for 1 hour at room temperature (18-24°C) in the dark.
  8. Aspirate and wash wells 5x with 300 µl diluted WASHBUF (Wash buffer). After final wash, remove remaining WASHBUF by strongly tapping plate against paper towel.
  9. Add 200 µl SUB (Substrate, blue cap) into each well.
  10. Incubate for 30 min at room temperature (18-24°C) in the dark.
  11. Add 50 µl STOP (Stop solution, white cap) into each well.
  12. Measure absorbance immediately at 450 nm with reference 630 nm, if available.

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Expected Values

Values from apparently healthy individuals:

  • Median Serum (n=411): 24.14 pmol/l

This value lies between calibration point 2 and 3 of the standard curve. It is recommended to establish the normal range for each laboratory.

Package Inserts

Please note: All documents above are for reference use only and should not be used in place of the documents included with this physical product. If digital copies are needed, please contact us.