Mouse Osteopontin DuoSet ELISA

Catalog # Availability Size / Price Qty
DY441
Ancillary Products Available
Mouse Osteopontin / OPN ELISA Standard Curve
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Product Details
Procedure
Citations (9)
FAQs
Supplemental Products
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Mouse Osteopontin DuoSet ELISA Summary

Assay Type
Solid Phase Sandwich ELISA
Format
96-well strip plate
Sample Volume Required
100 µL
Sufficient Materials
For fifteen 96-well plates*
Specificity
Please see the product datasheet

* Provided that the recommended microplates, buffers, diluents, substrates and solutions are used, and the assay is run as summarized in the Assay Procedure provided.

This DuoSet ELISA Development kit contains the basic components required for the development of sandwich ELISAs to measure natural and recombinant mouse Osteopontin (OPN). The suggested diluent is suitable for the analysis of most cell culture supernate samples. Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet.

Product Features

  • Optimized capture and detection antibody pairings with recommended concentrations save lengthy development time
  • Development protocols are provided to guide further assay optimization
  • Assay can be customized to your specific needs
  • Economical alternative to complete kits

Kit Content

  • Capture Antibody
  • Detection Antibody
  • Recombinant Standard
  • Streptavidin conjugated to horseradish-peroxidase (Streptavidin-HRP)

Other Reagents Required

DuoSet Ancillary Reagent Kit 2 (5 plates): (Catalog # DY008) containing 96 well microplates, plate sealers, substrate solution, stop solution, plate coating buffer (PBS), wash buffer, and Reagent Diluent Concentrate 2.

The components listed above may be purchased separately:

PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2PO4, pH 7.2 - 7.4, 0.2 µm filtered

Wash Buffer: (Catalog # WA126), or 0.05% Tween® 20 in PBS, pH 7.2-7.4

Reagent Diluent: (Catalog # DY995), or 1% BSA in PBS, pH 7.2-7.4, 0.2 µm filtered

Substrate Solution: 1:1 mixture of Color Reagent A (H2O2) and Color Reagent B (Tetramethylbenzidine) (Catalog # DY999)

Stop Solution: 2 N H2SO4 (Catalog # DY994)

Microplates: R&D Systems (Catalog # DY990)

Plate Sealers: ELISA Plate Sealers (Catalog # DY992)

Data Example

Mouse Osteopontin / OPN ELISA Standard Curve

Product Datasheets

Preparation and Storage

Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: Osteopontin/OPN

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.

Long Name:
Secreted Phosphoprotein 1 [BNSP]
Entrez Gene IDs:
6696 (Human); 20750 (Mouse); 25353 (Rat); 281499 (Bovine)
Alternate Names:
BNSP; Bone sialoprotein 1; Eta-1; MGC110940; Nephropontin; OPN; Osteopontin; secreted phosphoprotein 1bone sialoprotein I, early T-lymphocyteactivation 1); secreted phosphoprotein-1 (osteopontin, bone sialoprotein); Spp1; SPP-1; SPP1/CALPHA1 fusion; Urinary stone protein; uropontin

Assay Procedure

GENERAL ELISA PROTOCOL

Plate Preparation

  1. Dilute the Capture Antibody to the working concentration in PBS without carrier protein. Immediately coat a 96-well microplate with 100 μL per well of the diluted Capture Antibody. Seal the plate and incubate overnight at room temperature.
  2. Aspirate each well and wash with Wash Buffer, repeating the process two times for a total of three washes. Wash by filling each well with Wash Buffer (400 μL) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each step is essential for good performance. After the last wash, remove any remaining Wash Buffer by aspirating or by inverting the plate and blotting it against clean paper towels.
  3. Block plates by adding 300 μL Reagent Diluent to each well. Incubate at room temperature for a minimum of 1 hour.
  4. Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.

Assay Procedure

  1. Add 100 μL of sample or standards in Reagent Diluent, or an appropriate diluent, per well. Cover with an adhesive strip and incubate 2 hours at room temperature.
  2. Repeat the aspiration/wash as in step 2 of Plate Preparation.
  3. Add 100 μL of the Detection Antibody, diluted in Reagent Diluent, to each well. Cover with a new adhesive strip and incubate 2 hours at room temperature.
  4. Repeat the aspiration/wash as in step 2 of Plate Preparation.
  5. Add 100 μL of the working dilution of Streptavidin-HRP to each well. Cover the plate and incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
  6. Repeat the aspiration/wash as in step 2.
  7. Add 100 μL of Substrate Solution to each well. Incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
  8. Add 50 μL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
  9. Determine the optical density of each well immediately, using a microplate reader set to 450 nm. If wavelength correction is available, set to 540 nm or 570 nm. If wavelength correction is not available, subtract readings at 540 nm or 570 nm from the readings at 450 nm. This subtraction will correct for optical imperfections in the plate. Readings made directly at 450 nm without correction may be higher and less accurate.

Citations for Mouse Osteopontin DuoSet ELISA

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

9 Citations: Showing 1 - 9
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  1. OC-STAMP promotes osteoclast fusion for pathogenic bone resorption in periodontitis via up-regulation of permissive fusogen CD9
    Authors: T Ishii, M Ruiz-Torru, A Ikeda, S Shindo, A Movila, H Mawardi, A Albassam, RA Kayal, AA Al-Dharrab, K Egashira, W Wisitrasam, K Yamamoto, AI Mira, K Sueishi, X Han, MA Taubman, T Miyamoto, T Kawai
    FASEB J., 2018;0(0):fj201701424R.
    Species: Mouse
    Sample Types: Tissue Homoegenates
  2. Contribution of engineered nanomaterials physicochemical properties to mast cell degranulation
    Authors: MM Johnson, R Mendoza, AJ Raghavendr, R Podila, JM Brown
    Sci Rep, 2017;7(0):43570.
    Species: Mouse
    Sample Types: Cell Culture Supernates
  3. Physiologic Thymic Involution Underlies Age-Dependent Accumulation of Senescence-Associated CD4(+) T Cells
    Authors: K Sato, A Kato, M Sekai, Y Hamazaki, N Minato
    J. Immunol., 2017;0(0):.
    Species: Mouse
    Sample Types: Cell Culture Supernates
  4. Silica-Triggered Autoimmunity in Lupus-Prone Mice Blocked by Docosahexaenoic Acid Consumption
    PLoS ONE, 2016;11(8):e0160622.
    Species: Mouse
    Sample Types: BALF
  5. The impairment of osteogenesis in bone sialoprotein (BSP) knockout calvaria cell cultures is cell density dependent.
    Authors: Bouet G, Bouleftour W, Juignet L, Linossier M, Thomas M, Vanden-Bossche A, Aubin J, Vico L, Marchat D, Malaval L
    PLoS ONE, 2015;10(2):e0117402.
    Species: Mouse
    Sample Types: Cell Culture Supernates
  6. Skeletal development of mice lacking bone sialoprotein (BSP)--impairment of long bone growth and progressive establishment of high trabecular bone mass.
    Authors: Bouleftour W, Boudiffa M, Wade-Gueye N, Bouet G, Cardelli M, Laroche N, Vanden-Bossche A, Thomas M, Bonnelye E, Aubin J, Vico L, Lafage-Proust M, Malaval L
    PLoS ONE, 2014;9(5):e95144.
    Species: Mouse
    Sample Types: Serum
  7. Rosiglitazone improves survival and hastens recovery from pancreatic inflammation in obese mice.
    Authors: Pini M, Rhodes DH, Castellanos KJ
    PLoS ONE, 2012;7(7):e40944.
    Species: Mouse
    Sample Types: Plasma
  8. Vitamin D receptor agonists increase klotho and osteopontin while decreasing aortic calcification in mice with chronic kidney disease fed a high phosphate diet.
    Authors: Lau, Wei Ling, Leaf, Elizabet, Hu, Ming Cha, Takeno, Marc M, Kuro-o, Makoto, Moe, Orson W, Giachelli, Cecilia
    Kidney Int, 2012;82(12):1261-70.
    Species: Mouse
    Sample Types: Serum
  9. Phosphate feeding induces arterial medial calcification in uremic mice: role of serum phosphorus, fibroblast growth factor-23, and osteopontin.
    Authors: El-Abbadi MM, Pai AS, Leaf EM, Yang HY, Bartley BA, Quan KK, Ingalls CM, Liao HW, Giachelli CM
    Kidney Int., 2009;75(12):1297-307.
    Species: Mouse
    Sample Types: Serum

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