Figure 3

Sorted Nkx6.1⁺ progenitors that immediately differentiate in vitro generate the correct ventral neuronal subtypes. (A) Schematic diagram of the fluorescence-activated cell sorting (FACS), isolation and culture of Nkx6.1⁺ spinal progenitors. (B-G) Immunohistochemical analysis of neurons that differentiate in vitro without proliferation using Chx10 (V2a interneurons; B-C); Isl1/2 (MNs; D-E); and Nkx2.2 (V3 interneurons; F-G) antibodies. Tuj1 stains the neuron-specific βIII tubulin. (H, I) No Lim1/2⁺ neurons were present in cultures from sorted eGFP⁺ cells, although these neurons were born from sorted eGFP-negative progenitors (J, K). (L) Frequencies of three different neuronal subtypes generated in vitro from sorted Nkx6.1⁺ precursors. Bars represent mean ± s.e.m (n = 16 wells from 3 experiments).