293T cells
(3 × 105 cells) were plated in 12-well plates in triplicate
24 h before
transfection. An amount of 500 ng of the biosensor constructs was
transfected using PolyJet transfection reagent (SignaGen Laboratories).
After 48 h, supernatant or cells were collected. Cells were lysed
using passive lysis buffer (Promega), and NanoLuc luciferase assays
were performed using one of two substrates: furimazine, FMZ(Nano-Glo
Cell Reagent, Promega) or native coelenterazine, CTZ (3.33 uM final
concentration) (Nanolight Technologies – Prolume Ltd., Pinetop,
AZ, USA). Synergy Microplate Reader (BioTek, Winooski, VT, USA) was
used to measure luminescence. Results are presented as RLU (Relative
Luminescence Unit) normalized to control. The data presented are the
mean of three independent experiments.47 (link)
(3 × 105 cells) were plated in 12-well plates in triplicate
24 h before
transfection. An amount of 500 ng of the biosensor constructs was
transfected using PolyJet transfection reagent (SignaGen Laboratories).
After 48 h, supernatant or cells were collected. Cells were lysed
using passive lysis buffer (Promega), and NanoLuc luciferase assays
were performed using one of two substrates: furimazine, FMZ(Nano-Glo
Cell Reagent, Promega) or native coelenterazine, CTZ (3.33 uM final
concentration) (Nanolight Technologies – Prolume Ltd., Pinetop,
AZ, USA). Synergy Microplate Reader (BioTek, Winooski, VT, USA) was
used to measure luminescence. Results are presented as RLU (Relative
Luminescence Unit) normalized to control. The data presented are the
mean of three independent experiments.47 (link)
