Natural and recombinant mouse and rat OPN. This assay cross-reacts 8.2% with recombinant Human Osteopontin.
< 0.5% cross-reactivity observed with available related molecules.Cross-species reactivity observed with 1 or more species tested.
No significant interference observed with available related molecules.
The Quantikine Mouse OPN Immunoassay is a 4.5 hour solid-phase ELISA designed to measure mouse OPN in cell culture supernates, serum, plasma, and urine. It contains NS0-expressed recombinant mouse OPN and antibodies raised against the recombinant factor. This immunoassay has been shown to accurately quantitate the recombinant factor. Results obtained using natural mouse OPN showed linear curves that were parallel to the standard curves obtained using the Quantikine kit standards. These results indicate that this kit can be used to determine relative mass values for naturally occurring mouse OPN.
Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested on one plate to assess intra-assay precision.
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in separate assays to assess inter-assay precision.
The recovery of mouse OPN spiked to three levels throughout the range of the assay was evaluated.
Average % Recovery
Mouse Cell Culture Supernates (n=7)
Rat Cell Culture Supernates (n=7)
To assess the linearity of the assay, samples containing mouse or rat OPN were diluted with Calibrator Diluent and then assayed.
Preparation and Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Osteopontin (OPN), also known as bone sialoprotein (BSP), is a secreted SIBLING family protein that can be variably modified by O- and N-glycosylation, sulfation, phosphorylation, and transglutamination. OPN is widely expressed and is prominent in mineralized tissues. It inhibits bone mineralization and kidney stone formation and promotes inflammation, cell ad¬hesion, and migration. Its expression is upregulat¬ed during inflammation, obesity, atherosclerosis, cancer, and tissue damage and contributes to the pathophysiology of these conditions. The central region of OPN contains RGD and non-RGD binding sites for multiple integrins. Adjacent to the RGD motif is the sequence SLAYGLR (SVVYGLR in human) which serves as a cryptic binding site for additional integrins: it is masked in full length OPN but is exposed following OPN cleavage by multiple proteases in tumors and sites of tissue injury.
Refer to the product for complete assay procedure.
Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.
Prepare all reagents, standard dilutions, and samples as directed in the product insert.
Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.
50 µL Assay Diluent
Add 50 µL of Assay Diluent to each well.
50 µL Standard, Control, or Sample
Add 50 µL of Standard, Control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.
Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.
100 µL Conjugate
Add 100 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.
Aspirate and wash 5 times.
100 µL Substrate Solution
Add 100 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes. PROTECT FROM LIGHT.
100 µL Stop Solution
Add 100 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.
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