Fig. 3: Validation of the PECAn pipeline.
a Schematic of semi-synthetic image validation strategy. Z-planes with known parameters were randomly combined into Z-stacks. Known images were analysed using the macro and results were compared against known values. b Macro results divided by known values (n = 5 independent samples), with mean and 95% CI shown. No statistical test performed. All values approximate expected values, with deviations attributable to rounding errors. c Representative image of a competing wing disc stained with anti-cleaved Dcp-1. RpS3+/− patches are marked by GFP (green), and nuclei by DAPI staining (blue) (Left). The anti-cleaved Dcp-1 staining is shown (red) and the dotted line represents the outline of the loser patches (Right). d Comparison of the density of apoptotic events in the patch border as determined by hand against macro outputs. e Sample output of individual cell counts performed by PECAn in competing wing disc. Individual cells are marked with a uniquely colour-coded stamp. f Comparison of quantifications generated manually against those generated by the macro. g Representative images of wing discs harbouring competing RpS3+/− cells (green) immuno-stained for cleaved-Dcp-1 (red). From left to right, the genotypes are: no transgene other than GFP, expression of an empty UAS promoter, expression of an RNAi against Dronc, and expression of an RNAi against Xrp1. h Automated quantification of density of cleaved-Dcp-1-positive cells in the patch border region. Measure of center and error bars are shown as mean and 95% CI. Statistics reflect two-sided Wilcoxon–Mann–Whitney U-test without adjustment for multiple comparisons with Cliff’s δ effect size. Number of independent biological samples are: replicate 1: ncontrol = 6, nUAS = 6, nDronc = 4, nXrp1 = 6; replicate 2: ncontrol = 12, nDronc = 15. i Automated quantification of the percentage of the wing disc pouch occupied by RpS3+/− cells with mean and 95% CI shown. Statistics reflect two-sided t-test without adjustment for multiple comparisons with un-pooled Cohen’s d effect size. Number of independent biological samples samples per replicate are: replicate 1: ncontrol = 5, nUAS = 6, nDronc = 4, nXrp1 = 6; replicate 2: ncontrol = 12, nDronc = 15. Scale bars correspond to 50 µm. Source data are provided as a Source Data file.
