Description
Overview
Type I interferons, interferon alpha (IFN-α) and interferon beta (IFN-β), play essential roles in innate immune response to viral infections. IFN-β is produced by fibroblasts, macrophages, and dendritic cells, as well as endolial cells. expression of IFN-β gene is controlled by transcription factors like IRF-3, c-Jun/ATF-2, and NF-kappaB downstream of Pattern Recognition Receptors, such as Toll-like receptors. In particular, IFN-β is primary functional readout of cGas-cGAMP-STING pathway activation. Once secreted, IFN-β binds and activates IFNAR1/IFNAR2 receptor complex, which relies on STAT1/2 transcription factors. Phosphorylated STAT1/2 leads to induction of anti-viral gene expression.
Assessment of serum samples often requires enhanced sensitivity. In some cases, AlphaLISA assays may have sufficient sensitivity to enable detection of low levels of analytes in serum or plasma.
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.
