Mouse neurofibroma/DRG-derived sphere culture was performed as described (26). Specifically, we plated trypan blue–negative cells at 1 × 104 cells per 24-well low-binding plates in 1 ml of medium containing Dulbecco’s modified Eagle’s medium (DMEM):F-12 (3:1) and recombinant human epidermal growth factor (20 ng/ml; R&D Systems), 20 ng/ml recombinant human basic fibroblast growth factor (R&D Systems), 1% B-27 (Invitrogen), and heparin (2 μg/ml; Sigma-Aldrich). We maintained cultures at 37°C and 5% CO2. To passage, we centrifuged sphere cultures, dissociated and replated at 1 × 104 cells/ml in fresh sphere medium as described (6).

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