Despite the plethora of literature regarding isolation and characterization of periodontal ligament stem cells (PDLSCs), due to the existence of controversies in the results, in this comprehensive review, we aimed to summarize and compare the effect of isolation methods on PDLSC properties, including clonogenicity, viability/proliferation, markers expression, cell morphology, differentiation, and regeneration. Moreover, the outcomes of included studies, considering various parameters, such as teeth developmental stages, donor age, periodontal ligament health status, and part of the teeth root from which PDLSCs were derived, have been systematically discussed. It has been shown that from included studies, PDLSCs can be isolated from teeth at any developmental stages, health status condition, and donor age. Furthermore, a non-enzymatic digestion method, named as an explant or outgrowth technique, is a suitable protocol for PDLSCs isolation.
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