Dissociation and Single-Cell Analysis of Murine and Human Neuronal Cultures

Embryonic and neonatal C57/bl6 brains (n = 3 mice per time point) were dissociated in papain solution for 5 min at room temperature, followed by the addition of inhibitor solution and gentle trituration to generate single-cell suspensions. Human iPSC-derived neuronal cultures at day 45 of differentiation from TS patients and control subjects (n = 3 lines each for controls and patients) were dissociated in accutase, and FACS sorting was performed at the Stanford shared FACS facility as previously described (Paşca et al., 2011 (link); Ho et al., 2018 (link)), excluding debris, dead cells and doublets to sort single cells into multi-well plates. sc-qPCR experiments were performed as in previous studies (Paşca et al., 2011 (link); Yoo et al., 2011 (link); Portmann et al., 2014 (link)) on 96 × 96 dynamic arrays (Fluidigm) and as recommended by the array manufacturer.