Cytotoxicity Assay Protocol for Drugs

EC₅₀ was determined using AlamarBlue in 96-well plates [76 (link)]. All drugs were titrated in two-fold dilutions: Suramin from 1.56 to 400 nM, Pentamidine from 0.98 to 250 nM, Trypan Blue from 0.2 to 50 nM, Ponceau S from 0.39 to 100 nM and CiB from 0.98 to 250 nM. Drug exposure was for 72 hours and AlamarBlue incubation overnight. Plates were read on an Infinite 200Pro plate-reader (Tecan) with the following parameters: Excitation 530nm; emission 585 nm; filter cut-off 570 nm. Data were analysed in GraphPad PRISM. Asymmetric confidence intervals (95% CI) were calculated where possible and statistical significance of each EC₅₀ shift assessed by confidence interval overlap.