A new method to isolate microglia from adult mice and culture them for an extended period of time

As the major immuno-competent cells of the brain, microglia are highly implicated in neuro-protection as well as in neurodegeneration. Therefore, they are of key interest for research on numerous CNS diseases. Currently, to model inflammation in the brain, microglial cell lines or primary microglia prepared from embryonic or neo-natal rodents are widely used. However, these in vitro microglial models are not suitable for research in the field of neuro-degenerative diseases where aging is a crucial parameter. Only a few in vitro studies on aged microglia have been published so far, most of which use ex vivo microglia which cannot be kept in culture for prolonged periods of time. In the present study, we provide a new approach which allows the isolation and culture of an almost pure population of microglia from adult mouse brains. The isolation is based on a procedure which combines density separation and a subsequent culture selection process. After these steps, microglia form a non-adherent floating cell layer that can be easily and repeatedly harvested and replated. This method is simple and allows for a comparatively high yield and purity of adult microglial cells. The collected primary adult microglia proliferate and can be kept in culture for extended periods of time. We compared the primary adult microglia to primary microglia from neo-natal mice as well as to the C8-B4 microglial cell line. We found that adult microglia have similar, but not identical, immuno-phenotypic, functional and electrophysiological characteristics to the other in vitro models.