Preparation of the planarian Schmidtea mediterranea for high-resolution histology and transmission electron microscopy

The flatworm Schmidtea mediterranea is an emerging model species in fields such as stem cell biology, regeneration and evolutionary biology. Excellent molecular tools have been developed for S. mediterranea, but ultrastructural techniques have received far less attention. Processing specimens for histology and transmission electron microscopy (TEM) is notoriously idiosyncratic for particular species or specimen types. Unfortunately, however, most methods for S. mediterranea described in the literature lack numerous essential details, and those few that do provide them rely on specialized equipment that may not be readily available. Here we present an optimized protocol for ultrastructural preparation of S. mediterranea. The protocol can be completed in 6 d, much of which is 'hands-off' time. To aid with troubleshooting, we also illustrate the major effects of seemingly minor variations in fixative, buffer concentration and dehydration steps. This procedure will be useful for all planarian researchers, particularly those with relatively little experience in tissue processing.