Oligodendrocyte progenitor cells (OPCs) play a pivotal role in both health and disease within the central nervous system, with oligodendrocytes, arising from resident OPCs, being the main myelinating cell type. Disruption in OPC numbers can lead to various deleterious health defects. Numerous studies have described techniques for isolating OPCs to obtain a better understanding of this cell type and to open doors for potential treatments of injury and disease. However, the techniques used in the majority of these studies involve several steps and are time consuming, with current culture protocols using serum and embryonic or postnatal cortical tissue as a source of isolation. We present a primary culture method for the direct isolation of functional adult rat OPCs, identified by neuron-glial antigen 2 (NG2) and platelet derived growth factor receptor alpha (PDGFrα) expression, which can be obtained from the adult spinal cord. Our method uses a simple serum-free cocktail of 3 growth factors - FGF2, PDGFAA, and IGF-I, to expand adult rat OPCs in vitro to 96% purity. Cultured cells can be expanded for at least 10 passages with very little manipulation and without losing their phenotypic progenitor cell properties, as shown by immunocytochemistry and RT-PCR. Cultured adult rat OPCs also maintain their ability to differentiate into GalC positive cells when incubated with factors known to stimulate their differentiation. This new isolation method provides a new source of easily accessible adult stem cells and a powerful tool for their expansion in vitro for studies aimed at central nervous system repair.

Institute of Neuroscience Université catholique de Louvain Avenue Hippocrate B1 54 10 1200 Brussels Belgium

Louvain Drug Research Institute Advanced Drug Delivery and Biomaterials Université catholique de Louvain Avenue Mounier 73 B1 73 12 1200 Brussels Belgium

Louvain Drug Research Institute Advanced Drug Delivery and Biomaterials Université catholique de Louvain Avenue Mounier 73 B1 73 12 1200 Brussels Belgium Institute of Condensed Matter and Nanosciences Université catholique de Louvain 1348 Louvain la Neuve Belgium

Louvain Drug Research Institute Advanced Drug Delivery and Biomaterials Université catholique de Louvain Avenue Mounier 73 B1 73 12 1200 Brussels Belgium Integrated Center for Cell Therapy and Regenerative Medicine International Clinical Research Center St Anne's University Hospital Brno Pekařská 53 656 91 Brno Czech Republic

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