Flat clear bottom white polystyrene poly d lysine coated microplates

First, 10,000 iTF-iPSCs were seeded into 96-well Flat Clear Bottom White Polystyrene Poly-d-Lysine Coated Microplates (Corning; Cat. No. 3843) and differentiated into iTF-Microglia. For CSF1R inhibition, cells were treated with the CSF1R inhibitor PLX3397 at day 8 (ApexBio; Cat. No. B5854) at indicated concentrations or dimethylsulfoxide control for 24 h before performing the CellTiter-Glo 2.0 (Promega; Cat. No. G9242) assay according to the manufacturer’s instructions. For MAPK14 inhibition, cells were treated with the MAPK14 inhibitor Skepinone-L at day 8 (Selleckchem; Cat. No. 1221485831) at indicated concentrations or dimethylsulfoxide control for 24 h before performing the CellTiter-Glo 2.0 assay. Luminescence signal was recorded with the M5 plate reader (SpectroMax) using SoftMax Pro 6.5.1.

First, 10,000 iTF-iPSCs were seeded into 96-well Flat Clear Bottom White Polystyrene Poly-d-Lysine Coated Microplates (Corning; Cat. No. 3843) and differentiated into iTF-Microglia as described above. At day 8 of the differentiation, the medium of iTF-Microglia was replaced with (1) full media change of iTF-Microglia medium containing 2 μg ml⁻¹ doxycycline, or (2) full media change of iTF-Microglia medium containing no doxycycline, or (3) half media changes of iTF-Microglia medium containing no doxycycline. This media-replacing paradigm was repeated every 3 d until day 15. Microglia survival was assessed by performing the CellTiter-Glo 2.0 (Promega; Cat. No. G9242) assay according to the manufacturer’s instructions. Luminescence signal was recorded with the M5 plate reader (SpectroMax).