1450 microbeta trilux luminescence microplate reader

Ultra-low attachment, round-bottomed 96-well plates (Corning Costar) were used for spheroid formation. SW480, SW480-CLDN1 or SW620 cells were seeded at a density of 5 × 10⁴. Cells aggregated and merged in 3D spheroids within 24–72 h. Images of wells were taken with a phase-contrast microscope using a 5× objective or captured with the Celigo™ imaging cytometer using the “Tumorosphere” application. Cell viability was assessed with the CellTiter-Glo Luminescent Cell Viability Assay (Promega, Madison, WI, USA). After addition of 100 μl of CellTiter Glo reagent to each well for 10 min, luminescence was measured on a 1450 MicroBeta TriLux Luminescence microplate reader (Perkin Elmer).

Ultra-low attachment, round-bottomed 96-well plates (Corning Costar) were used for spheroid formation. HCT116 cells were seeded at a density of 150 cells/well. Cells aggregated and merged to form 3D spheroids within 24-72 h. Spheroids were then grown in the presence of BKM120 (75, 150, 300, 600 and 1200 nM) and MEK162 (0.3, 1, 3, 10, 30, 90, 270 nM) (added at day 4 and day 10 of culture) and experiments ended 14 days after seeding. Images of wells were taken with a phase-contrast microscope using a 10X objective. Cell viability was assessed with the CellTiter-Glo Luminescent Cell Viability Assay (Promega, Madison, WI, USA). After addition of 100 μl of CellTiter Glo reagent to each well for 10 min, luminescence was measured on a 1450 MicroBeta TriLux Luminescence microplate reader (Perkin Elmer).