ADAR1:RNA TR-FRET Assay Kit
The ADAR1:RNA TR-FRET Assay Kit is designed to measure the binding of ADAR1 (adenosine deaminase, RNA-specific 1) to RNA for screening and profiling applications using TR-FRET (Time-Resolved Fluorescence Resonance Energy Transfer). It utilizes Terbium-labeled donor and dye-labeled acceptor to complete the TR-FRET pairing. The ADAR1:RNA TR-FRET Assay Kit comes in a convenient 384-well format, with enough purified ADAR1, Tb-Labeled Donor and Dye-Labeled Acceptor, RNase inhibitor, ADAR1 substrate and assay buffer for 384 reactions. The assay also includes a Competitor RNA as control.
Figure 1: ADAR1 mechanism of action on RNA molecules.
Figure 2: Assay principle.
The Terbium-labeled anti-FLAG antibody donor binds to FLAG-tagged ADAR1, while the dye-labeled streptavidin acceptor binds to the biotinylated RNA substrate. When ADAR1 forms a complex with its RNA substrate TR-FRET occurs and can be measured using a TR-FRET fluorescence plate reader. TR-FRET signal is proportional to RNA binding.
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- Fluorescent microplate reader capable of measuring Time Resolved Fluorescence Resonance Energy Transfer (TR-FRET)
- Incubator set to 30°C
- Nuclease-free water
- Adjustable micropipettor and sterile filter tips
- Rotating or rocker platform
| Catalog # | Name | Amount | Storage |
| 100472 | ADAR1, FLAG-Tag* | 3 µg | -80°C |
| ADAR1 Substrate, Biotin-Labeled (10 µM) |
20 µl | -80°C | |
| RNase Inhibitor | 100 µl | -80°C | |
| Competitor RNA (10 µM) | 10 µl | -80°C | |
| 3x ADAR Binding Buffer | 3 ml | -20°C | |
| Anti-FLAG Tb-Labeled Donor | 2 x 10 µl | -20°C | |
| Dye-Labeled Acceptor | 10 µl | -20°C | |
| 79969 | White 384-well microplate | 1 | Room Temp |
*The initial concentration of enzyme is lot-specific and will be indicated on the tube containing the protein.
ADAR1 (adenosine deaminase, RNA-specific 1) performs adenosine to inosine base editing in RNA, particularly targeting adenosines located within a specific stem-loop motif structure. It is proposed that ADARs evolved to provide additional diversity to the transcriptome and while the majority of ADAR editing events occur in non-coding RNAs, some, including the canonical GluA2 editing site, alter the amino acid sequence of coding proteins. ADAR1 plays a role in innate immunity by mitigating interferon signaling. Dysfunction of ADAR1 results in autoimmune disorders, and impacts cancer cell growth and proliferation as well as tumor response to immunotherapy. Since ADAR recognizes double-stranded RNA, it also functions to suppress or modify RNA viruses. Thus, it is implicated in viral evolution and in the emergence of viral variants such as SARS-CoV-2 variants.
Keegan L. P., et al., 2023 Acc Chem Res 56 (22): 3165-3174.
Mendoza H. G., et al., 2023 Biochemistry 62 (8): 1367-1387.
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