Fig 4. Repeatability, reproducibility and robustness of the HCI assay.

A-Plot of the number of host cells in each well of a set of 3 96-well plates to calculate the intra-plate and inter-plate variation. HFFs incubated with DMSO were represented with black dots while those incubated with pyrimethamine were represented with white squares. B- Plot of Toxoplasma proliferation to assess the robustness. HFF incubated with pyrimethamine (white squares) or DMSO (black dots). The Z’ factor was also calculated. C-Pyrimethamine dose-response curve and determination of the IC₅₀ at 3 different days to calculate reproductibility. HFFs were incubated with Toxoplasma parasites and pyrimethamine. Vacuoles and parasites were counted to calculate the parasite proliferation inhibition and to determine the IC₅₀.