Keratinocyte stem cell colonies can be identified by measuring cell locomotion speed. (A) Clonal analysis of human clonogenic keratinocytes (Barrandon and Green, 1987b). A progressive growing colony derived from a single keratinocyte is subcultured into a 10-cm cell culture dish (indicator dish) so its growing capacity can be evaluated. (B, top) Progressive growing colonies showed variations in the mean cell locomotion speed in the colony after 7 d of cultivation. The mean cell locomotion speed in the colony was obtained by measuring locomotion speed of randomly selected 15–20 cells in each colony. Bar, 100 µm. (bottom) Each growing colony in the top panel was trypsinized and passaged into an indicator dish arranged in the same column. The culture was maintained for 12 d, and then fixed and stained with rhodamine B. Proliferative capacity of growing colonies was assessed by determining the ratio of terminal colonies in the indicator dish. Bars, 10 mm. (C) Correlation of mean speed of cells in each colony and its proliferative capacity (Pearson’s r = −0.477, P = 0.0335, and n = 20 colonies). (D) Growing colonies were categorized in three groups depending on the mean speed of cells: slow (≤30 µm/h), medium (>30 µm/h and ≤35 µm/h), and fast (>35 µm/h). P-values were calculated by Student’s t test.