Glomax multiplus plate reader luminometer

ATP content was evaluated using Cell Titer-Glo® 3D Cell Viability Assay (Promega) according to manufacturer’s instructions. Briefly, 50 μl of the reagent was added to individual spheroids in 100 μl of culture medium. To facilitated lysis, organoids were vortexed for 1 min and the plate was incubated at 37°C in 5% CO₂ for 20 min with subsequent luminescent signal measurement using GloMax® Multiplus Plate Reader/Luminometer (Promega). The changes in viability are represented as % compared to viability of control spheroids/organoids. The viability of organoids after 48 h of incubation with tested compounds was visualized using a LIVE/DEAD® assay (Thermo Fisher Scientific) as described by the manufacturer. Briefly, organoids were washed in DPBS and incubated for 30 min at 37°C in a 5 % CO₂ incubator in 1 mL of culture media containing 1 μL of calcein AM solution and 5 μL of ethidium homodimer-1 solution. Stained spheroids/organoids were analyzed using a fluorescence microscope (Zeiss).

Cell viability of VeroE6/TMPRSS2 was measured by an MTS [3-(4,5-dimethylthylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay using CellTiter 96 AQueous One Solution (Promega, Madison, WI, USA) in the presence of different concentration of CPC (0–50 µg/mL) for 1 h at 37 °C. The absorbance was measured with GloMax Multiplus Plate Reader/Luminometer (Promega). Three independent experiments were performed in triplicate.