Fig 1. Schematic representation of culture schedule and characterization of IVMDE.

(A). Schematic representation of culture schedules for rat/mouse IVMDE in comparison to the typical in vitro myelination protocol using rat dissociated neurons. (B) MBP mRNA expression levels in rat IVMDE after initiation of myelination by ascorbic acid. MBP expression levels at indicated days after initiation of myelination are shown normalized to GAPDH expression relative to the expression on day 2. The data from each experiment are shown as S1 File. (C-E) Representative immunocytochemistry photomicrographs of rat IVMDE at 14 days after induction of myelination by ascorbic acid to detect βⅢ-tubulin depicted in green, and MBP depicted in red are shown. Scale bar = 1mm. Insets show high-power view of the rectangular area in each panel. Arrows in (D) indicate nodes of Ranvier identifiable in light microscopy. (F, G) Representative immunocytochemistry photomicrographs of MBP in IVMDE (F) and conventional dissociated culture (G), both using rat cells. Scale bar = 50μm. (H) The number of myelin profiles in in vitro myelination culture using dissociated neurons and in IVMDE are shown relative to the number in dissociated neuron experiments. The myelination profiles were counted in 5 microscopic fields randomly selected from 5 DRG cultures (1 field per each DRG culture) in 3 independent culture experiments. *p<0.01. The data from each experiment are shown as S1 File. (I~L) Representative photomicrographs for EM analysis of rat IVMDE culture showing myelinated (I, J) and non-myelinated (K, L) axons/SCs are shown. Scale bar = 1μm (I, K, L), 100 nm (J).