Table 3 The procedure and reagents used for differentiation of hEnSCs into motor neuron-like cells on 3D collagen hydrogels with or without EpoB-loaded PCL microspheres or cell culture plates.
Step | Differentiation medium | Ingredients | Catalog number | Duration |
|---|---|---|---|---|
1 | Pre-induction (PrI) medium | DMEM/F12 (1:1) | Invitrogen, 32500-035 | 1 days |
20% FBS | Gibco, 10270-106 | |||
2% B-27 supplement | Gibco, 17504-044 | |||
100 μM 2-Mercaptoethanol | Sigma, M3148 | |||
250 μM isobutylmethylxanthine (IBME) | Sigma, I5879 | |||
10 ng/mL fibroblast growth factor 2 (FGF2) | Sigma, SRP4037 | |||
2 | Induction medium (exchanged after 3 days) | DMEM/F12 (1:1) | Invitrogen, 32500-035 | 7 days |
0.2% B-27 supplement | Gibco 17504-044 | |||
100 ng/mL sonic hedgehog (SHH) | Sigma, SRP3156 | |||
0.01 μM retinoic acid (RA) | Sigma, R2625 | |||
3 | Maturation (survival) medium (exchanged after 3 days) | DMEM/F12 (1:1) | Invitrogen, 32500-035 | 7 days |
0.2% B-27 supplement | Gibco 17504-044 | |||
100 ng/mL glial cell-derived neurotrophic factor (GDNF) | Sigma, G1777 | |||
200 ng/mL brain-derived neurotrophic factor (BDNF) | Sigma, SRP3014 |
