Supplementary Figure 4: Enrichment of postmitotic hMNs by FACS.
From: ALS-implicated protein TDP-43 sustains levels of STMN2, a mediator of motor neuron growth and repair
a, Using a cell surface marker screen, we identified antibodies enriched on GFP+ motor neurons (Quadrant 1) and GFP- cells (Quadrant 3). b, After sorting for NCAM+ and EpCAM- cells, we used high content imaging to determine our sorting method can deplete the cultures of mitotic cells (EdU+) and significantly enrich for motor neurons (Isl1+) and neurons (MAP2+). n = 6 different iPSC lines (unpaired t-test, two-sided, P < 0.05). c–e, qRT-PCR analysis of sorted and unsorted cultures for the neuronal marker βIII-tubulin (c) and the motor neuron markers ISL1 (d) and VaCHT1 (e) revealed enrichment after sorting for NCAM+ and EpCAM- cells. Data are displayed as mean of technical replicates with s.d. from n = 6 iPSC lines (unpaired t-test, two-sided, P < 0.05). f, Flow-cytometric analysis with phycoerythrin (PE)-conjugated antibodies to EpCAM (anti-epCAM–PE) and Alexa Fluor 700–conjugated antibodies to NCAM (anti-NCAM–AF700) of cultures differentiated from the indicated healthy controls (gray) and TDP-43 mutant lines (red). g, The percentage of NCAM+ cells for the indicated lines. Data are displayed as mean with s.d. from n = 4 independent differentiations (unpaired t-test, two-sided, P < 0.05). h–i, NCAM+ neurons from 11a were plated onto multielectrode arrays to measure neuronal activities including spiking and bursting over time. Similar results were obtained for n = 3 iPSC lines sorted using NCAM.