The development of new imaging techniques offers the possibility to go beyond the analysis of mostly flat surfaces and image and analyze complex tissue organization in depth. Here, we describe a method, called DISSECT, that offers the opportunity to extract automatically, in strongly deformed epithelia, a precise characterization of the spatial organization of a given cytoskeletal network combined with morphological quantifications in highly remodeled three-dimensional (3D) epithelial tissues. We believe that this method, applied here to Drosophila tissues, will be of general interest in the expanding field of morphogenesis and tissue biomechanics.