Fractionation of hematopoietic progenitors from differentiated cells in OP9/ES cell differentiation system. (A) Formation of cobblestone is observed with a phase-contrast microscopy at day 14 of ES cell culture. Dark round cells indicated by arrows are hematopoietic progenitors lying beneath OP9 layer. Large irregular-shaped cells indicated by arrowheads are OP9 cells. (B) Schematic presentation of the method to prepare floating and adherent fractions. First, floating fractions are harvested with culture media. Adherent cells are then disaggregated transiently by trypsin and transferred to a new culture dish. After a 1-hour incubation, which makes OP9 cells adhere to the dish, the floating cells are obtained from media (adherent fraction).