Fig. 4

Reproducible miniaturization of organoid cultures to 5 µl without detectable growth deficits. A Multiway ANOVA table of P values for organoid size and the indicated fixed effects and interaction terms. Culture volumes (2 µl, 5 µl, and 7 µl) were considered together and in pairs (n = 2–6 factors or levels per effect). Specific illustrations of fixed effects are shown in subsequent subpanels. B Differences in organoid size distribution for four representative patient sources. 5 µl organoid cultures were measured for cross-sectional area by brightfield imaging on day 7. C Differences in organoid size distribution at 2, 5, 7, 10, 12 and 14 days. The 5 µl organoid culture was measured longitudinally as in (B). D Differences in organoid size distribution between 2 µl, 5 µl, and 7 µl cultures. The patient-derived sample was measured on day 7 as in (B). E Overlapping variance of downsampled technical replicates among organoid culture volumes. Organoid counts in the larger volumes of three patients were repeatedly downsampled to the 2 µl median (n = 1000 iterations; tan), and variance was estimated after shifted log transformation (see Methods). Variance estimates of the entire organoid count from each volume and technical replicate are overlaid (black). See Additional file 3: Fig. S9 for analysis of additional patients. For (B–D), size distributions are summarized by their cumulative distribution function