For in vitro myelination assay, O4+ RRMS and control hiOL were purified by flow cytometry at day 21 of differentiation. Afterwards, 1 × 105 O4+ hiOL were seeded in DM onto aligned nanofibers (Nanofiber solutions) that were coated with 1 µg/mL PLO and 5 µg/mL Laminin LN211 (Biolamina) beforehand. For staining of nanofibers, coating was performed by replacing PLO with 100 µg/mL PLL-FITC (Sigma). Half of the medium was changed every other day. After 7 days, cells were fixed and stained for MBP. A previously published heuristic algorithm was used for quantification [66 (link)].
Starost L., Lindner M., Herold M., Xu Y.K., Drexler H.C., Heß K., Ehrlich M., Ottoboni L., Ruffini F., Stehling M., Röpke A., Thomas C., Schöler H.R., Antel J., Winkler J., Martino G., Klotz L, & Kuhlmann T. (2020). Extrinsic immune cell-derived, but not intrinsic oligodendroglial factors contribute to oligodendroglial differentiation block in multiple sclerosis. Acta Neuropathologica, 140(5), 715-736.
Other organizations :
University Hospital Münster, Johns Hopkins Medicine, Johns Hopkins University, Max Planck Institute for Molecular Biomedicine, Vita-Salute San Raffaele University, Istituti di Ricovero e Cura a Carattere Scientifico, University of Münster, Montreal Neurological Institute and Hospital, McGill University, Friedrich-Alexander-Universität Erlangen-Nürnberg, Universitätsklinikum Erlangen
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