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HTRF Mouse IFN-β Detection Kit, 500 Assay Points
HTRF Mouse IFN-β Detection Kit, 500 Assay Points
Generic HTRF biomarker image
| Feature | Specification |
|---|---|
| Application | Protein Quantification |
| Sample Volume | 16 µL |
Product variants
Unit Size: 500 assay points
Part #:
62MIFNBPEG
Unit Size: 10,000 assay points
Part #:
62MIFNBPEH
For research use only. Not for use in diagnostic procedures. All products to be used in accordance with applicable laws and regulations including without limitation, consumption, and disposal requirements under European REACH regulations (EC 1907/2006).
HTRF Mouse IFN-β Detection Kit, 500 Assay Points
Generic HTRF biomarker image
HTRF Mouse IFN-β Detection Kit, 500 Assay Points
Product information
Overview
Type I interferons, interferon alpha (IFN-α) and interferon beta (IFN-β), play essential roles in the innate immune response to viral infections. IFN-β is produced by fibroblasts, macrophages, and dendritic cells, as well as endothelial cells. The expression of the IFN-β gene is controlled by transcription factors like IRF-3, c-Jun/ATF-2, and NF-kappaB downstream of Pattern Recognition Receptors, for example Toll-like receptors. In particular, IFN-β is the primary functional readout of cGas-cGAMP-STING pathway activation. Once secreted, IFN-β binds and activates the IFNAR1/IFNAR2 receptor complex, which relies on STAT1/2 transcription factors. Phosphorylated STAT1/2 leads to the induction of anti-viral gene expression.
How it works
Assay principle
Cell supernatant, sample, or standard is dispensed directly into the assay plate for the detection by HTRF® reagents (384-well low-volume white plate or Revvity low-volume 96-well plate in 20 µL). The antibodies labeled with the HTRF donor and acceptor are pre-mixed and added in a single dispensing step, to further streamline the assay procedure. The assay can be run up to a 1536-well format by simply resizing each addition volume proportionally.
Assay data analysis
The 4 Parameter Logistic (4PL) curve is commonly recommended for fitting an ELISA standard curve. This regression enables the accurate measurement of an unknown sample across a wider range of concentrations than linear analysis, making it ideally suited to the analysis of biological systems like cytokine releases.
To fully understand how to deal with HTRF data processing and also 4PL 1/y² fitting,
Revvity has also worked with Myassays.com to help you with your data analysis. you’ll be able to access a free online software to run your IFN beta analysis.
Assay details
Technical specifications of mouse IFN beta kit
| Sample size | 16 µL |
|---|---|
| Final assay volume | 20 µL |
| Kit components | Lyophilized standard, frozen detection antibodies, buffers, & protocol |
| LOD & LOQ (in Diluent) | 4 pg/mL & 17 pg/mL |
| Range | 17 - 2,000 pg/mL |
| Time to result | Overnight at RT |
| Species | Mouse only |
Analytical performance
Intra and inter assay
Intra assay (n=24)
| Sample | Mean [IFN-β] (pg/mL) | CV |
|---|---|---|
| 1 | 75 | 10% |
| 2 | 200 | 3% |
| 3 | 1125 | 3% |
| Mean CV | 5.3% |
Inter assay (n=6)
| Sample | Mean [IFN-β] (pg/mL) | CV |
|---|---|---|
| 1 | 58 | 3.6% |
| 2 | 348 | 3.3% |
| 3 | 1092 | 7.3% |
| Mean CV | 4.7% |
Assay validation
The mouse STING-specific agonists CMA and DMXAA trigger IFN-β release from NIH-3T3 fibroblast cells
The mouse NIH-3T3 fibroblast cell line was plated in a 96-well culture plate (100,000 cells/well) and incubated for 6h. The cells were then treated with 200 µM of the synthetic mouse STING-specific agonists CMA and DMXAA (under 100 µL). After 24h incubation, cell supernatants were collected and secreted levels of IFN-beta were measured following the procedure given in the HTRF kit’s package insert.
IFN-β secretion by the murine RAW 264.7 macrophage cell line treated with several STING agonists
The murine RAW 264.7 macrophage cell line was plated in a 96-well culture plate (100,000 cells/well) and incubated overnight. The cells were then treated with a fixed concentration of the synthetic STING agonists CMA, DMXAA, and ADUS100, as well as with the endogenous STING agonist 2'3'-cGAMP (under 100 µL). After 24h secretion, cell supernatants were collected and levels of released IFN-beta were measured following the procedure given in the HTRF kit’s package insert.
Dose-dependent secretion of IFN-β by the murine RAW 264.7 macrophage cell line treated with 2’3’-cGAMP and ADUS100
The murine RAW 264.7 macrophage cell line was plated in a 96-well culture plate (100,000 cells/well) and incubated for 5h. The cells were then treated with increasing concentrations of either the endogenous STING agonist 2'3'-cGAMP, or the synthetic small molecule ADUS100 (under 100 µL). After an overnight secretion, cell supernatants were collected and levels of released IFN-beta were measured following the procedure given in the HTRF kit’s package insert.
Specifications
| Application |
Protein Quantification
|
|---|---|
| Brand |
HTRF
|
| Detection Modality |
HTRF
|
| Product Group |
Kit
|
| Sample Volume |
16 µL
|
| Shipping Conditions |
Shipped in Dry Ice
|
| Target Class |
Cytokines
|
| Target Species |
Mouse
|
| Technology |
TR-FRET
|
| Therapeutic Area |
Oncology & Inflammation
|
| Unit Size |
500 assay points
|
Video gallery
HTRF Mouse IFN-β Detection Kit, 500 Assay Points
HTRF Mouse IFN-β Detection Kit, 500 Assay Points
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