Human embryonic stem cells' (hESCs) unlimited proliferative potential and differentiation capability to all somatic cell types made them potential cell source in different cell-based tissue engineering strategies as well as various experimental applications in fields such as developmental biology, pharmacokinetics, toxicology, and genetics. Periodontal tissue engineering aims to improve the outcome of regenerative therapies which have variable success rates when contemporary techniques are used. Cell-based therapies may offer potential advantage in overcoming the inherent limitations associated with guided tissue-regeneration procedures, such as dependency on defect type and size and the pool and capacity of progenitor cells resident in the wound area. Elucidation of developmental mechanisms of different periodontal tissues may also contribute to valuable knowledge based upon which the future therapies can be designed. Prior to the realization of such a potential, protocols for the differentiation of pluripotent hESCs into periodontal ligament fibroblastic cells (PDLF) as common progenitors for ligament, cementum, and alveolar bone tissue need to be developed. The present protocol describes methods associated with the guided differentiation of hESCs by the use of coculture with adult PDLFs, and the resulting change of morphotype and phenotype of the pluripotent embryonic stem cells toward fibroblastic and osteoblastic lineages.