For each kinase, 10 µL of transfection ready working DNA from plates A and B was transferred into 96-well assay plates (Corning #3917), with one well corresponding to a top control well and one to a sample well. This was followed by addition of Fugene HD transfection reagent (Promega #E2311), 30 µL of 20 µL/mL stock in each well containing DNA. Assay plates were mixed on an orbital shaker for 15 seconds at 300 rpm and incubated at room temperature for 30 min. HEK293 cells at 2.5 x 10 5 cells/mL in Opti-MEM without phenol red (Gibco #11058-021) containing 1% FBS (Avantor Seradigm #97068-085) were added to the assay plate (60 µL per well) and incubated for 20-24 hours at 37°C and 5% CO2. Prior to tracer and inhibitor addition, the assay plates were equilibrated at room temperature for 15 min.