Recent advances in tridimensional (3D) tissue imaging and development of reporter proteins have provided new insight into the dynamic relationship between tissue structure, function and dysfunction in a number of fields of research. In developmental biology 3D microscopy has enabled building of quantitative 3D atlases of embryo morphology and gene expression at cellular resolution and in neuroscience connectomics, the building of 3D brain connectivity maps is a rapidly expanding area of research. Recently, in endocrinology, in vivo whole tissue 3D imaging by the use of two photon excitation microscopy has revealed the importance of cellular networks in the pituitary and endocrine pancreas which direct development and function. However, optical microscopy methods are limited in resolution compared to electron microscopy. Advances in electron microscopy include the development of serial block face scanning electron microscopy which enables the 3D visualisation of fixed tissues with fine ultrastructural detail. Furthermore, correlative light and electron microscopy (CLEM) integrates the advantages of 3D light and electron microscopy on the same sample to enable the subcellular localisation of dynamic processes in vivo with the resolution power of the electron microscope. Most recently the emerging technology of light-sheet microscopy allows live imaging of organs with high spatiotemporal resolution over long periods of time. The current state of the art in 3D biological imaging techniques will be explored with a focus on recent applications to the study of endocrine systems.