Flow cytometry methods for the study of cell-cycle parameters of planarian stem cells

Due to their characteristic inaccessibility and low numbers, little is known about the cell-cycle dynamics of most stem cells in vivo. A powerful, established methodology to study cell-cycle dynamics is flow cytometry, which is used routinely to study the cell-cycle dynamics of proliferating cells in vitro. Its use in heterogeneous mixtures of cells obtained from whole animals, however, is complicated by the relatively low abundance of cycling to non-cycling cells. We report on flow cytometric methods that take advantage of the abundance of proliferating stem cells in the planarian Schmidtea mediterranea. The optimized protocols allow us to measure cell-cycle dynamics and follow BrdU-labeled cells specifically in complex mixtures of cells. These methods expand on the growing toolkit being developed to study stem cell biology in planarians, and open the door to detailed cytometric studies of a collectively totipotent population of adult stem cells in vivo.