A novel method for isolating Schwann cells using the extracellular domain of Necl1.
Myelinating cocultures of Schwann cells and dorsal root ganglion neurons are a powerful experimental system for probing the molecular mechanisms of axon-Schwann cell interaction. The isolation of a pure population of myelination-competent Schwann cells is a prerequisite for this experimental system. We describe here a protocol for a FACS-based isolation of Schwann cells utilizing a specific affinity reagent (Necl1-Fc) and the use of these isolated cells in myelinating cocultures. An advantage of the myelinating coculture system is that Schwann cells and the neurons can be genetically manipulated before they are cocultured. We further show that our method allows the isolation of virally transduced Schwann cells in a single purification step. This protocol for the FACS-based isolation of myelination-competent Schwann cells by Necl1-Fc and the use of these cells in myelinating cocultures should significantly facilitate future studies aimed at delineation of the molecular mechanisms of axon-Schwann cell interactions and myelination.
Spiegel
Ivo
I
Department of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot, Israel.
Peles
Elior
E
D000818:Animals; D015816:Cell Adhesion Molecules, Neuronal / Q000737:chemistry* / Q000378:metabolism; D002450:Cell Communication / Q000502:physiology; D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002478:Cells, Cultured; D018920:Coculture Techniques; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000378:metabolism; D005434:Flow Cytometry / Q000379:methods*; D005727:Ganglia, Spinal / Q000166:cytology / Q000378:metabolism; D005822:Genetic Vectors / Q000494:pharmacology; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D009488:Neurosciences / Q000379:methods*; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000378:metabolism; D012583:Schwann Cells / Q000166:cytology / Q000378:metabolism*; D014161:Transduction, Genetic / Q000379:methods
D015816:Cell Adhesion Molecules, Neuronal; D017077:Culture Media, Conditioned; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; C516997:nectin-like molecule 1, rat