SOST, also known as Sclerostin, is a Cerberus/DAN family member and is an important regulator of bone homesotasis. Cerberus/DAN proteins (Cerberus, DAN, Gremlin, PRDC, and Caronte) are secreted glycoproteins that function as BMP antagonists. While the overall sequence identity between members of the family is low, they share a cysteine-knot motif with conserved spacing of six cysteine residues. SOST is secreted as a monomer in contrast to many other cysteine-knot proteins which form disulfide-linked homodimers. Mature mouse SOST shares 89% and 95% aa sequence identity with human and rat SOST, respectively. Inactivating mutations in the SOST gene can cause sclerosteosis and van Buchem disease which are bone dysplasia disorders characterized by progressive skeletal overgrowth. SOST is expressed by terminally differentiated cells embedded in mineralized matrix including osteocytes, hypertrophic and prehypertrophic chondrocytes, and tooth cementocytes. SOST expression is induced by BMP-2, -4, and -6 and is inhibited by parathyroid hormone (PTH). Its downregulation in osteocytes by physical loading of bone contributes to the mechanical sensor function of osteocytes and the subsequent increase in bone growth. SOST binds to BMP-2, -4, -5, -6, and -7 and inhibits the alkaline phosphatase (ALP) activity induced by these BMPs. It inhibits canonical Wnt signaling by binding to LRP-5 and LRP-6 and inhibiting their association with Frizzled receptors. SOST also modulates the ability of BMPR-IA signaling to interfere with canonical Wnt signaling. These interactions underlie the ability of SOST to inhibit BMP- and Wnt-induced bone formation in vivo. SOST reduces the proliferation of mesenchymal stem cells (MSC) and induces MSC apoptosis. It inhibits the differentiation of preosteoblastic cells and bone mineralization by osteoblasts. In knockout mice that lack SOST expression, osteocyte and osteoblast apoptosis is inhibited, osteoblast activity is enhanced, and the mice are resistant to mechanical unloading-induced bone loss. SOST knockout mice also exhibit increased bone mineral density, bone volume, and bone strength throughout the skeleton in trabecular and cortical bone. Mice treated with neutralizing SOST antibodies likewise show increased bone formation and bone mineral density. This treatment can reverse the bone loss and bone integrity decline that is otherwise seen in models of osteoporosis and chronic gut inflammation. The SOST, BioAssay™ ELISA Kit is a 4.5h solid-phase ELISA designed to measure SOST in mouse or rat cell culture supernates, serum, and plasma. It contains NS0-expressed recombinant mouse SOST and antibodies raised against the recombinant factor. This immunoassay has been shown to accurately quantitate the recombinant mouse SOST. Results obtained using natural mouse or rat SOST showed dose response curves that were parallel to the standard curves obtained using the kit standards. These results indicate that this kit can be used to determine relative mass values for natural SOST.
Sample Type:Cell culture supernates, serum, and plasma
Intended Use:For the quantitative determination of mouse or rat SOST concentrations in cell culture supernates, serum, and plasma.
Sensitivity:~1.63pg/ml
Range:0.422-4.17pg/ml
Specificity:Mouse or rat SOST
Test Principle:This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for mouse/rat SOST has been pre-coated onto a microplate. Standards, control, and samples are pipetted into the wells and any SOST present is bound by the immobilized antibody. After washing away any unbound substances, an enzyme-linked polyclonal antibody specific for mouse/rat SOST is added to the wells. Following a wash to remove any unbound antibody-enzyme reagent, a substrate solution is added to the wells. The enzyme reaction yields a blue product that turns yellow when the Stop Solution is added. The intensity of the color measured is in proportion to the amount of SOST bound in the initial step. The sample values are then read off the standard curve.
Kit Components:SOST Microplate: 96 well polystyrene microplate (12 strips x 8 wells) coated with a monoclonal antibody specific for mouse/rat SOST
SOST Conjugate: 1x12ml of polyclonal antibody specific for mouse/rat SOST conjugated to horseradish peroxidase with preservatives
SOST Standard: 5000 pg of recombinant mouse SOST in a buffered protein base with preservatives; lyophilized
SOST Control: 1 vial of recombinant mouse SOST in a buffered protein base with preservatives; lyophilized. The concentration range of SOST after reconstitution is shown on the vial label. The assay value of the Control should be within the range specified on the label
Assay Diluent RD1W: 1x12ml of a buffer with preservatives
Calibrator Diluent RD6-12: 1x21ml of a buffered protein base with preservatives
Wash Buffer Concentrate: 1x50ml of a 25-fold concentrated solution of buffered surfactant with preservativesColor Reagent A: 1x12ml of stabilized hydrogen peroxideColor Reagent B: 1x12ml of stabilized chromogen (tetramethylbenzidine)Stop Solution: 1x23ml of diluted hydrochloric acidPlate Sealers: 4 adhesive strips
Storage and Stability:See Kit Protocol for detailed storage instructions.
