Total TBK1 cellular kit
All-in-one kit for robust detection of Total TBK1
The human RIG-I binding assay is designed to select and characterize compounds that specifically bind human RIG-I protein.
A fast and easy way to identify new binders to human RIG-I. The Retinoic acid-Inducible Gene I (RIG-I) is a cytosolic pattern recognition receptor which is known to recognize RNA pathogens in cells. This protein is involved in the innate immune system and induces a type 1 interferon response. The RIG-I protein recognizes infected cells by detecting viral dsRNA. Once activated by a dsRNA, the N-terminus domain of RIG-I binds mitochondrial antiviral signaling proteins, thereby activating the IFN-1 signaling pathway.
The HTRF RIG-I binding assay is a competitive assay format which uses a biotinylated 3 phosphate-dsRNA, a streptavidin-d2, a 6His tagged human RIG-I protein, and an anti 6His Cryptate-labeled antibody. The compound being tested competes with the biotinylated 3p-dsRNA, and thereby prevents FRET from occurring.
The Human RIG-I binding assay can be run in a 96- or 384-well low volume white plate (20 µL final). As described here, samples or standards are dispensed directly into the assay plate. The human His-tagged RIG-I protein is then added, followed by the dispensing of the HTRF reagents: The anti 6His antibody labeled with Terbium cryptate and the biotinylated-3p-dsRNA bound to d2-labeled streptavidin. The reagents labeled with HTRF fluorophores may be pre-mixed and added in a single dispensing step. No washing steps are needed. The protocol can be further miniaturized or upscaled by simply resizing each addition volume proportionally.
Two different ds-RNAs were tested in parallel in the HTRF RIG-I binding assay. The first ds-RNA displays a triphosphate in 5', the second does not. Serial dilutions of both RNAs were carried out in diluent, and then RIG-I protein was dispensed. HTRF detection reagents were then added to the well for the final detection step. Results were collected after 1 hour of incubation at room temperature. The affinity of the RIG-I protein was increased by the presence of the triphosphate in 5' of the dsRNA, as expected.
Best practices for pharmacological characterization of PPI inhibitors
Easy pharmacological characterization of PPI modulators. - Technical Notes
HTRF assays handle low- to high affinity protein-protein interactions
Deciphering low- and high affinity interactions - Application Notes
Nuclear receptor ligand identification with HTRF
Monitoring nuclear receptor binding with HTRF assays - Application Notes
HTRF addresses large protein-protein interaction complexes
Challenge large complexes with HTRF assays - Application Notes
Virology research solutions using HTRF Protein-Protein Interaction assays
See how peer researchers challenge the viral life cycle with PPI assays - Application Notes
Product Insert Bioch. RIGI (h) Kit / 64BDRIGIPEG-64BDRIGIPEH
64BDRIGIPEG-64BDRIGIPEH - Product Insert
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