| Assay Method Information | |
| | Bioluminescence Resonance Energy Transfer (BRET) Assay |
| Description: | The inhibitory activity of each compound on the binding between NRF2 and KEAP1 was measured by bioluminescence resonance energy transfer (BRET) assay. A plasmid having human NRF2 (Original Clone ID: ha01449s1) sequence fused with NanoLuc (19 kDa, Promega) at the N-terminal was transduced into HCE-T cells (immortalized human corneal epithelial cells) using FuGENE HD (Promega). 48 hours after transduction, the cell lysate was collected and used as an NRF2-expressing cell lysate. A plasmid having human KEAP1 (Original Clone ID: cs00328) sequence fused with HaloTag (33 kDa, Promega) at the C-terminal was transduced into HCE-T cells using FuGENE HD (Promega). After 24 hours, HaloTag Nano BRET 618 Ligand (Promega) was added to the medium. The cell lysate cultured for 24 hours after the addition was used as a KEAP1-expressing cell lysate. |
| Affinity data for this assay | |
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