Production of Highly Uniform Midbrain Organoids from Human Pluripotent Stem Cells

Figure 1

Generation of µMOs using a high-throughput platform. (a) Schematic illustration of the procedure for generating µMOs. The procedures and timelines for generating µMOs and typical MOs are described. (b) The morphology and quality of MOs generated from a different number of starting cells (DIV 14 and 21). Scale bar, 1 mm. (c) Average diameter of MOs generated from a different number of starting cells on DIV 1, 7, 14, and 21 of differentiation. Data are presented as mean ± SD (n = 20 for each number of starting cells). (d) Expression of forebrain (FOXG1 and LHX2), midbrain (LMX1B and TH), and hindbrain (HA1 and HB4) specific marker genes in MOs generated from a different number of starting cells was analyzed by qPCR (DIV 14). Expression levels are normalized to those of undifferentiated hESCs. Typical cerebral organoids (COs), MOs, and hindbrain organoids (HOs) were also used for positive controls. Data are presented as mean ± SD of triplicate values. (e and f) Representative confocal images of MOs generated from a different number of starting cells (DIV 21). Scale bar, 100 μm. (g) Representative confocal images of MOs generated from a different number of starting cells expressing TH (DIV 30). Scale bar, 100 μm. (h) Percentage of TH-positive cells in µMOs generated from a different number of starting cells (DIV 30). Data are presented as mean ± SD of triplicate values. (i) The representative section images displaying global enrichment of MAP2 and TH positive cells in µMOs (DIV 30). Scale bar, 100 μm.

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