Description
Mouse OPG (Osteoprotegerin) ELISA Kit
The Mouse OPG (Osteoprotegerin) ELISA Kit is a reliable tool for the accurate measurement of OPG levels in mouse serum, plasma, and cell culture supernatants. With its high sensitivity and specificity, this kit provides consistent and precise results, making it suitable for various research purposes. Osteoprotegerin is a key regulator of bone metabolism, acting as a decoy receptor for RANKL and inhibiting osteoclast differentiation and bone resorption.
Dysregulation of OPG has been linked to various bone-related diseases, including osteoporosis and bone metastasis, highlighting its importance as a biomarker for studying these conditions and potentially developing therapeutic interventions. Whether investigating bone health, cancer metastasis, or other related research areas, the Mouse OPG ELISA Kit offers a valuable tool for accurate and efficient analysis of Osteoprotegerin levels in mouse samples.
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Mouse |
| Detection Method: | Colormetric |
| Detection Range: | 62.50-4000 pg/mL |
| Sensitivity: | 37.50 pg/mL |
| Sample Volume Required Per Well: | 100µL |
| Sample Type: | Serum, plasma and other biological fluids |
| Specificity: | This kit recognizes Mouse OPG in samples. No significant cross-reactivity or interference between Mouse OPG and analogues was observed. |
This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Mouse OPG. Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Mouse OPG and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Mouse OPG, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Mouse OPG. The concentration of Mouse OPG in samples can be calculated by comparing the OD of the samples to the standard curve.
| UniProt Protein Function: | TNFRSF11B: Acts as decoy receptor for RANKL and thereby neutralizes its function in osteoclastogenesis. Inhibits the activation of osteoclasts and promotes osteoclast apoptosis in vitro. Bone homeostasis seems to depend on the local RANKL/OPG ratio. May also play a role in preventing arterial calcification. May act as decoy receptor for TRAIL and protect against apoptosis. TRAIL binding blocks the inhibition of osteoclastogenesis. Defects in TNFRSF11B are the cause of juvenile Paget disease (JPD); also known as hyperostosis corticalis deformans juvenilis or hereditary hyperphosphatasia or chronic congenital idiopathic hyperphosphatasia. JPD is a rare autosomal recessive osteopathy that presents in infancy or early childhood. The disorder is characterized by rapidly remodeling woven bone, osteopenia, debilitating fractures, and deformities due to a markedly accelerated rate of bone remodeling throughout the skeleton. Approximately 40 cases of JPD have been reported worldwide. Unless it is treated with drugs that block osteoclast- mediated skeletal resorption, the disease can be fatal. |
| UniProt Protein Details: | Protein type:Secreted; Inhibitor; Secreted, signal peptide Cellular Component: proteinaceous extracellular matrix; extracellular space; extracellular region Biological Process: extracellular matrix organization and biogenesis; apoptosis; negative regulation of odontogenesis of dentine-containing teeth; negative regulation of bone resorption; signal transduction |
| UniProt Code: | O08712 |
| NCBI GenInfo Identifier: | 21264085 |
| NCBI Gene ID: | 18383 |
| NCBI Accession: | O08712. 1 |
| UniProt Secondary Accession: | O08712,O70202, |
| UniProt Related Accession: | O08712 |
| Molecular Weight: | 45,923 Da |
| NCBI Full Name: | Tumor necrosis factor receptor superfamily member 11B |
| NCBI Synonym Full Names: | tumor necrosis factor receptor superfamily, member 11b (osteoprotegerin) |
| NCBI Official Symbol: | Tnfrsf11b |
| NCBI Official Synonym Symbols: | Opg; TR1; OCIF |
| NCBI Protein Information: | tumor necrosis factor receptor superfamily member 11B; osteoprotegerin; osteoclastogenesis inhibitory factor |
| UniProt Protein Name: | Tumor necrosis factor receptor superfamily member 11B |
| UniProt Synonym Protein Names: | Osteoclastogenesis inhibitory factor; Osteoprotegerin |
| Protein Family: | Tumor necrosis factor receptor superfamily |
| UniProt Gene Name: | Tnfrsf11b |
| UniProt Entry Name: | TR11B_MOUSE |
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e. g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
| Concentration (pg/mL) | O.D | Average | Corrected |
| 4000 | 2.286 2.326 | 2.306 | 2.25 |
| 2000 | 1.468 1.488 | 1.478 | 1.422 |
| 1000 | 0.938 0.908 | 0.923 | 0.867 |
| 500 | 0.452 0.466 | 0.459 | 0.403 |
| 250 | 0.227 0.213 | 0.22 | 0.164 |
| 125 | 0.161 0.141 | 0.151 | 0.095 |
| 62.50 | 0.1 0.11 | 0.105 | 0.049 |
| 0 | 0.052 0.06 | 0.056 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Mouse OPG were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Mouse OPG were tested on 3 different plates, 20 replicates in each plate.
| Intra-assay Precision | Inter-assay Precision | |||||
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean (pg/mL) | 217.50 | 502.90 | 1434.50 | 232.90 | 462.30 | 1300.20 |
| Standard deviation | 14.10 | 25.60 | 77.50 | 11.90 | 26.40 | 58.50 |
| C V (%) | 6.48 | 5.09 | 5.40 | 5.11 | 5.71 | 4.50 |
Recovery
The recovery of Mouse OPG spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
| Sample Type | Range (%) | Average Recovery (%) |
| Serum (n=5) | 93-106 | 99 |
| EDTA plasma (n=5) | 89-103 | 94 |
| Cell culture media (n=5) | 87-101 | 93 |
Linearity
Samples were spiked with high concentrations of Mouse OPG and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
| Serum (n=5) | EDTA plasma (n=5) | Cell culture media (n=5) | ||
| 1:2 | Range (%) | 92-106 | 87-100 | 90-100 |
| Average (%) | 99 | 92 | 95 | |
| 1:4 | Range (%) | 96-111 | 84-96 | 95-107 |
| Average (%) | 102 | 91 | 100 | |
| 1:8 | Range (%) | 96-112 | 84-98 | 97-112 |
| Average (%) | 102 | 89 | 104 | |
| 1:16 | Range (%) | 100-114 | 85-97 | 94-109 |
| Average (%) | 107 | 90 | 102 |
An unopened kit can be stored at 4°C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
| Item | Specifications | Storage |
| Micro ELISA Plate(Dismountable) | 8 wells ×12 strips | -20°C, 6 months |
| Reference Standard | 2 vials | |
| Concentrated Biotinylated Detection Ab (100×) | 1 vial, 120 µL | |
| Concentrated HRP Conjugate (100×) | 1 vial, 120 µL | -20°C(shading light), 6 months |
| Reference Standard & Sample Diluent | 1 vial, 20 mL | 4°C, 6 months |
| Biotinylated Detection Ab Diluent | 1 vial, 14 mL | |
| HRP Conjugate Diluent | 1 vial, 14 mL | |
| Concentrated Wash Buffer (25×) | 1 vial, 30 mL | |
| Substrate Reagent | 1 vial, 10 mL | 4°C(shading light) |
| Stop Solution | 1 vial, 10 mL | 4°C |
| Plate Sealer | 5 pieces | |
| Product Description | 1 copy | |
| Certificate of Analysis | 1 copy |
- Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- Aliquot 100µl of standard solutions into the standard wells.
- Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix. Incubate for 1 hour at 37°C.
- Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- Add 90µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37°C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
- Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.
