I have seen many papers and discussions here about various techniques for isolation and culture of microglia and astrocytes from adult mouse brain. Isolation of these cells by Miltenyi magnetic beads is supposed to yield a great purity and have been used for short-term culture (6-24h?) and RNA analysis. Can anyone give me feedback from their experience about separation of cells using Miltenyi beads and culturing these cells for a week (or at least up to 72h) , as described in the Miltenyi protocol?
The beads do work well! I concentrate the microglia using Percoll centrifugation and then label with the CD11b MACS antibody. The cultures last upwards of 2-3 weeks on a PDL coated substrate. ScienCell's microglia medium works well or DMEM-F12 with 10% FBS + 1%P/S is suitable for maintenance. Age of the mice is the most crucial aspect of getting high yields (see Article Isolation and Culture of Microglia
) . Best of luck!Elliot
Hi Elliot,
Thanks for your prompt response. Why poly D lysine...poly L lysine is not good to coat the plates?
I am trying to culture mixed glial from adult mouse 2-3 months old. I can only get microglia but not astrocytes. But when I culture from p0 mouse I can see astrocytes and microglia. I am using DMEM -f12 media with antibiotics. Using PDL coated dishes. Also I am using papain for digestion for the tissue. Does anybody have insight on this?
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