Fig. 1

Characterisation of undifferentiated and differentiated CTX0E16 hNPCs. a, b Representative images of undifferentiated (days differentiated (DD) 0) CTX0E16 cells immunostained for the neural progenitor cell (NPC) markers nestin (a) and Sox2 (b). c At DD 0, the majority of CTX0E16 NPCs were positive for the cell proliferation marker, KI67. d By DD 5, the number of KI67-positive CTX0E16 cells had greatly reduced. e Quantification of the number of KI67 positive cells as a percentage of total cells; n = approximately 1,500 cells from three independent experiments carried out in triplicate; error bars represent SD; ***p <0.001 (Student’s unpaired t test). f, g Only a few cells were positive for the astrocyte marker S100β at DD 0 (f) or DD 28 (g). h, i At DD 0 very few cells expressed the neuronal marker Tau (h). However, after 28 days of differentiation, the majority of cells expressed Tau (i). j, k Similarly, very few cells were positive for the neuronal marker MAP2 at DD 0 (j), but by DD 28 the majority of cells were positive for MAP2, indicating that the vast majority of cells at this time point had differentiated into neurons. l Number of proliferative (DD 0) or differentiated (DD 28) cells positive for S100β, Tau or MAP2; N = approximately 1,500 cells from three independent experiments carried out in triplicate; error bars represent SD; ***p <0.001 (two way analysis of variance with Tukey post hoc analysis). m Quantitative PCR (q-PCR) analysis revealed an increase in the expression of dorsal forebrain marker, EMX1 and cortical neuron marker TBR1 in DD 28 CTX0E16 neurons compared to DD 0 cells; n = 4 independent experiments carried out in triplicate; error bars represent SD; ***p <0.001 (Student’s unpaired t test). Scale bar, 50 μm. DAPI 4′,6-diamidino-2-phenylindole