A practical CD90.2-based FACS strategy for obtaining Schwann cell-enriched cultures from adult mouse nerves.

Primary Schwann cell cultures derived from adult murine peripheral nerves are frequently compromised by fibroblast overgrowth, limiting their utility for in vitro studies. Existing enrichment strategies often rely on selective growth conditions, prolonged culture, or antibody-mediated positive selection, which may alter Schwann cell physiology, trigger unwanted signaling, or result in incomplete fibroblast removal. To address these limitations, we developed a reproducible negative selection protocol for the isolation and enrichment of primary adult murine Schwann cells using fluorescence-activated cell sorting (FACS). By specifically targeting contaminating fibroblasts using the marker CD90.2 (Thy-1.2), these cells are efficiently depleted while leaving the Schwann cell population unbound by antibody complexes. This approach enables the rapid generation of highly enriched Schwann cell populations without the need for extended in vitro expansion or antimitotic agents. By avoiding direct receptor-ligand interactions on Schwann cells, this method preserves their native physiological state and surface epitopes, supporting sensitive downstream applications. Successful Schwann cell enrichment was confirmed by quantitative PCR, western blot and immunocytochemistry, demonstrating the enrichment of Schwann cell-specific markers and minimal fibroblast contamination. Overall, this approach enables the rapid generation of highly enriched Schwann cell populations while preserving features relevant to mature Schwann cell biology. This protocol provides a robust and practical tool for researchers requiring physiologically representative adult Schwann cells for molecular, functional, and co-culture-based studies of the peripheral nervous system.