The Kriks et al. protocol has finally achieved what we believe are genuine midbrain, dopamine neurons. Perrier et al. (2004) described a method to make dopamine neurons that we initially thought were mDA neurons. Later work by developmental biologists, however, discovered additional markers of these neurons that the Perrier cells lacked (e.g., FOXA2/TH).
Dual SMAD inhibition takes a confluent, feeder free culture of hPSCs and rapidly differentiates them into early neurectoderm. This rapid differentiation is caused by blocking the two signaling pathways that utilize SMADs for transduction: BMP and TGFB. Oct4 is extinguished and Pax6 expression has begun by around day 7–8, depending on the line used.