Twenty-five species of cetaceans have been reported throughout the Mediterranean Sea, eight of them are commonly distributed in the whole basin and are regularly found beached or adrift in the sea. Stranded animals are frequently found in poor conservation status, preventing reliable identification; identification is thus often based solely on morphological features. Therewith, molecular tools are especially useful to provide taxonomic identification. In this work, a four-enzymes PCR-RFLP in silico protocol, based on a fragment of the mitochondrial gene cytb, has been designed for cetacean species occurring in the Mediterranean Sea. Moreover, beached or floating specimen samples belonging to the eight common species have been tested in the laboratory, providing evidence that this approach represents a reliable, cost- and time-effective tool for their specific identification.