Fig. 5.

Establishment of a long-term cultured neurosphere line BXD-2664PNET-NS. (A) Images showing the morphology of neurospheres at different sizes. The cultures usually split when large neurospheres (arrow) were readily seen. (B) Profiles showing the FCM analysis of CD133⁺ and CD15⁺ subpopulations in the cultured neurospheres during serial passages through double staining. Neurospheres predigested with trypsin to dissociate into single cells were allowed to recover overnight before being double-stained with fluorochrome-conjugated antibodies specific to human CD133 and CD15. (C) Representative images showing the immunofluorescent staining of the cultured neurosphere cells. Following incubation with the primary antibodies against markers associated with stem cells (nestin), mature glial cells (GFAP), immature neuron (TuJ), oligodendrocyte (O4) and vimentin (VMT), fluorochrome-conjugated secondary antibodies (red for nestin, and green for the rest of the antibodies) were applied. The slides were counterstained with DAPI to highlight cell nuclears in blue. Original magnification: x 40.