Volume 18, Number 2, June 2006

Cultivation of Human Oral Keratinocytes and Fibroblasts Using an Explant Technique

Xue-Peng Xiong,1,2 San-Gang He,1,2 Jie-Mei Zhai,2 Ming Xu,2 Yi-Fang Zhao1
1Department of Oral and Maxillofacial Surgery, School and Hospital of Stomatology, Wuhan University, and
2Key Laboratory of Oral Biomedical Engineering, Wuhan University, Ministry of Education, Wuhan, China

To report a simple and reliable method for culturing normal human oral keratinocytes in a culture system free from feeder cells, with separation of the contaminating fibroblasts and their propagation.
Materials and Methods: The primary culture was initiated by explantation. The contaminating fibroblasts were isolated through trypsinization for their propagation. Subcultivation of oral keratinocytes by cell drops was done when 70 % to 80 % confluence was achieved. Morphological characteristics of both cell types were assessed under phase-contrast microscopy and the origin of the cultured cells was determined by cytohistochemical staining. The growth potential of both types of cells was monitored by growth curves.
Results: Oral keratinocytes could be serially cultured through 4 passages, and the contaminating fibroblasts were propagated after separation through trypsinisation. The morphology of the oral keratinocytes was typical pavement-like phenotype and was positive to cytokeratin staining. The growth curves of both cell types were S shaped.
Conclusion: This explant culture method may be a useful and economical option for serial cultivation of oral keratinocytes and fibroblasts.

Key words: Cell culture techniques, Fibroblasts, Immunohistochemistry, Keratinocytes, Mouth mucosa

Asian J Oral Maxillofac Surg. 2006;18:99-104.

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