Fig. S6

Enhanced apoptosis in the caudal fin-folds of Pten deficient embryos. (A) Uncut embryos from a ptena^+/-ptenb^-/- in-cross were fixed at 4dpf (4dpf, uncut) in parallel to the embryos depicted in Fig. 6A. (B, C) Embryos from a ptena^+/-ptenb^-/- in-cross were amputated and fixed at 1dpa (i.e. 3dpf, 1dpa) , equivalent uncut controls (3dpf, uncut) were included. Embryos were subjected to whole-mount immunohistochemistry using an antibody specific for the apoptosis marker activated Caspase-3 (green). The embryos were counterstained with DAPI (blue). Representative images of embryo caudal fin-folds are shown, and in the left panels the edge of the fin-fold is indicated with a dashed line. Number of embryos showing similar patterns/ total number of embryos analysed is indicated in the bottom right corner. The scale bar represents 100?m. Zoom-in images of each caudal fin-fold are shown. White arrowheads indicate cells; red arrowheads indicate background staining. (D) Caspase-3 immunofluorescence was quantified by mean particle count of the caudal fin-fold following thresholding and size restriction to reduce background signal. Means within amputated or uncut groups are compared to ptena^+/+ptenb^-/- embryos. The number of embryos analysed is indicated (n). Significance: * p<0.05; ** p<0.01; error bars represent standard deviation. Quantification of Caspase-3 immunofluorescence at 4dpf is depicted in Fig. 6B.