Skip to main content
. Author manuscript; available in PMC: 2020 Nov 12.
Published in final edited form as: J Vis Exp. 2019 Nov 12;(153):10.3791/60440. doi: 10.3791/60440

Figure 3: Representative sort plots of ventral midbrains (A) and ventral spinal cords (B).

Figure 3:

(Aa and Ba). Forward Scatter Area (FSC-A) versus Side Scatter Area (SSC-A) sort plot before exclusion of debris and dead cells. Ab, Bb, Ac, Bc) Sort plots for exclusion of cell clumps (b) and doublets (c) based on Width (SSC-W) versus SSC-A and Forward Scatter Width (FSC-W) versus FSC-A, respectively. Ad and Bd) Sort plots to isolate IslMN:GFP -positive motor neurons. Set the GFP gate higher for SMNs (Bd) than for CN3s/CN4s (Ad) in order to obtain a pure culture. Ae and Be) Percentages of cells gated for collection by FACS sorting. %Parent represents the percentage of cells in the current gated population relative to the number of cells in the previous gated cell population, whereas %Total represents the percentage of gated cells relative to total cells. Expected percentages of GFP-positive cells as compared to total cells (boxed in red) are 0.5–1.5 % for CN3/CN4 and 1.5–2.5% for SMN. If dissection was performed successfully, these percentages can be used as a benchmark to set up the GFP-positive gate in (Ad and Bd).