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Pan-cellular staining and variable-thickness views allow for characterization of 3D pathological features in wild-type larval and <italic>huli hutu</italic> mutant specimens.Top: Cutout visualization of both wild-type and huli hutu larval (five dpf) zebrafish stained with PTA showing detail in many soft tissue structures. Bottom: Cell types and structures that can be visualized include neuronal cells in the eye (A), cartilaginous rudiments of the squamous patch on the dorsal (arrow) pharynx (B), nucleated red blood cells (C), intact pneumatic duct (* to arrow) and goblet cells in the gut (D), and cross-striations of bands of muscles encircling the swim bladder (E). Panels A and D represent individual slices (0.743 μm in thickness) while B, C, E represent maximum intensity projections of 5 μm thick sections to visualize larger 3D structures. Compared to age matched wild-type larval zebrafish (top), the number of neuronal cells in the eye are markedly reduced (A’), chondrocytes appear cytologically normal, but formation of cartilaginous structures is markedly reduced (B’), the myocardium is thickened and, as is evident from a survey through all the sections of the heart (a single slice shown here) contains a markedly reduced number of nucleated red blood cells, consistent with anemia and abnormal hematopoiesis (C’). We are able determine the absence of the pneumatic duct and swim bladder in hht due to the ability to scan through the full volume of the sample. D’ shows degenerate tissue and E’ other tissues where those organs normally lie. A’, B’ and D’ represent individual slices (0.743 μm in thickness) while C’ and E’ represent maximum intensity projections of 5 μm thick sections to visualize structures of larger dimension.
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