Fig. 2: Testing a set of potential reference compounds with SPACe.

U-2 OS were treated for 24 h with the indicated compounds at 10 µM ahead of the CP protocol. Representative 20× color images from the same experiment out of three independent biological replicates are shown in (A). Scale bar is 10 µm. B Heatmap showing the signed EMD (Earth Mover’s Distance) from DMSO controls of the indicated compounds from three independent biological replicates (labeled in parentheses below the name of the compounds as 1,2,3). Each channel is separated to show the changes by fluorescent dye. N/A represents shape features and ratios (i.e., nuclear size/cell size). C Euclidean distance is used as a measure to compare the features fingerprint of each compound from the median DMSO control. This is measured for each well (represented as a hollow circle in the graph) treated with the indicated compound in three independent biological replicates, shown together in the graph. Overlayed in red the mean ± standard deviation is represented. * is p < 0.05 using non-parametric ANOVA (Kruskal–Wallis) compared to DMSO group. D Stacked bar graphs representing the fraction of changed features (in blue if they are reduced or red if they are increased, gray means no change) in the defined groups for each treatment. The chosen threshold for significance was EMD distance <−0.15 or >0.15 (see “Methods” section for description). Source data are provided as a Source Data file.