Abstract The purpose of this report is to present a case with dens invaginatus (DI) and to demonstrate the outcome of regenerative endodontic treatment (RET) with platelet rich fibrin (PRF) of the associated periapical lesion. DI is an enamel lined developmental malformation that occurs through an invagination of the dental papilla during the soft tissue stage of tooth development. Although often seen in maxillary lateral incisors, it very rarely occurs in mandibular premolar teeth; only 10 cases have been defined between 1993 and 2016. Its accurate diagnosis using 3D imaging methods is essential to examine the complex root canal structure and to prevent any associated caries, pulp necrosis, periapical lesions and periodontal problems. In the present case report, a mandibular right second premolar with DI and related periapical lesion was treated by using RET with PRF, which is defined as a biologically-based procedure to replace the damaged structures of the pulp-dentin complex and to restore the normal physiologic functions. A follow-up examination after 12 months showed that the tooth was free of any clinical symptoms, and its periapical condition was normal radiographically.