Figure 5. Generation and characterisation of hNMPs.

(A) Scheme describing the culture conditions employed for neural differentiation of hES cells treated for 72 h either with FGF/CHIR or subjected to dual SMAD inhibition (LDN, LDN193189; SB43, SB431542). (B) BRACHYURY/SOX2 immunocytochemistry in undifferentiated and FGF/CHIR-treated (48 h) hES cells. Corresponding graphs depict image analysis of BRACHYURY and SOX2 expression in the indicated culture conditions. Numbers: percentages of cells in each quadrant. (C) qPCR analysis for indicated markers in hES cells treated with FGF/CHIR for 72 h (D3) or 96 h (D4). Error bars = s.d. (n = 2). Results are represented as log₁₀ ratio of expression versus untreated hES cells. (D) qPCR analysis for indicated differentiation markers in hES cells differentiated in N2B27 following either an NM progenitor induction- (N_P) or a dual SMAD inhibition-intermediate step (N_A). Error bars = s.d. (n = 2). Anterior, anterior neural markers; PXM, paraxial mesoderm; n/d, not determined. (E) Immunocytochemistry for SOX2/HOXC8 in N_A and N_P culture conditions indicated in the scheme (A). (F) Quantitation of the coexpression of Hoxc8 with Sox2 in N_A and N_P conditions. All data used to generate the plots can be found in Data S5.