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Stem Cells International
Volume 2012, Article ID 485950, 15 pages
http://dx.doi.org/10.1155/2012/485950
Research Article

Clonal Populations of Amniotic Cells by Dilution and Direct Plating: Evidence for Hidden Diversity

1Wake Forest School of Medicine, Institute for Regenerative Medicine, Medical Center Blvd., Winston-Salem, NC 27157, USA
2Wake Forest University, Medical Center Blvd., Winston-Salem, NC 27157, USA
3Wake Forest School of Medicine, Medical Center Blvd., Winston-Salem, NC 27157, USA

Received 3 March 2012; Revised 9 May 2012; Accepted 29 May 2012

Academic Editor: Toshio Nikaido

Copyright © 2012 Patricia G. Wilson et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Fetal cells are widely considered a superior cell source for regenerative medicine; fetal cells show higher proliferative capacity and have undergone fewer replicative cycles that could generate spontaneous mutations. Fetal cells in amniotic fluid were among the first normal primary cells to be cultured ex vivo, but the undefined composition of amniotic fluid has hindered advance for regenerative applications. We first developed a highly efficient method to generate clonal populations by dilution of amniocentesis samples in media and direct plating without intervening refrigeration, centrifugation, or exposure of cells to the paracrine effects in mixed cell cultures. More than 40 clonal populations were recovered from 4 amniocentesis samples and representative clones were characterized by flow cytometry, conventional assays for differentiation potential, immunofluorescence imaging, and transcript analysis. The results revealed previously unreported diversity among stromal and epithelial cell types and identified unique cell types that could be lost or undetected in mixed cell populations. The differentiation potential of amniotic cells proved to be uncoupled from expression of definitive cell surface or cytoplasmic markers for stromal and epithelial cells. Evidence for diversity among stromal and epithelial cells in amniotic fluid bears on interpretations applied to molecular and functional tests of amniotic cell populations.