FIG. 3.

Sorting of neurosphere cells based on morphological characteristics and neurosphere-forming potential of the sorted populations. (A) FSC/SSC profile of passage 2 E14.5 neurosphere cells. FSC/SSC^high and FSC/SSC^low populations were defined based on FSC/SSC signal intensity. (B) FSC/SSC^high singlets and (C) FSC/SSC^low singlets were gated and sorted for the experiments to avoid doublets. (D) Percentage of FSC/SSC^high and FSC/SSC^low population based on total live-cell population (mean±SD; n=5; ***P≤0.001). (E) Neurosphere-forming potential expressed as NFU, which refers to the number of neurospheres formed per 100 cells plated. NFA was done at low density (1000 cells/mL) (mean±SD; n=5; ***P≤0.001) and (F) at clonal density (1 cell/well) after sorting cells into FSC/SSC^high and FSC/SSC^low populations (mean±SD; n=4; *P≤0.05). (G) Fold difference in neurosphere formation for FSC/SSC^high compared with FSC/SSC^low cells or unsorted cells at low and clonal density (mean±SD; n≥4). FSC, forward scatter; SSC, side scatter; E, embryonic day; NFU, neurosphere-forming unit; NFA, neurosphere formation assay; SD, standard deviation.