Analysis of proliferation and survival of DN3 and DN4 cells in the presence and absence of Notch signaling. (A) DN3 and DN4 cells derived from C57Bl/6 mice were labeled with CFSE and cocultured on OP9-DL1 monolayers with GSI (bold line) or without GSI (thin line) (1 μM). Proliferation was examined on day 7 by flow cytometric analysis of CFSE-labeled cells. (B) Cell counts for DN3 and DN4 cells derived from C57Bl/6 mice cocultured on OP9-DL1 monolayers with or without different concentrations of the GSI are shown. Cells were plated at an initial density of 5 × 10⁴ cells/well, cultured in the presence of DMSO or increasing concentrations of GSI, and analyzed after 7 days of coculture. Total cellularity is plotted as a function of GSI concentration.