Isolation of Schwann Cells from Mouse Spinal Roots
Transcription
Take a nerve root and chop it into small pieces.
Transfer to a digestive enzyme-containing tube and incubate.
The enzyme digests the extracellular matrix.
Add media containing serum to stop digestion.
Pipette the content repeatedly, then centrifuge and discard the supernatant.
Resuspend the pellet.
Pass the suspension through a strainer to remove undigested tissue. Incubate the collected cells in a plate.
Fibroblast and Schwann cells or SCs attach to the plate and divide.
Remove the unattached cells. Wash the attached cells with a phosphate buffer.
Add digestive enzymes to loosen the digestion-sensitive SCs. Stop digestion with media.
Completely detach SCs by adding media.
Transfer the cells to a tube and centrifuge. Discard the supernatant and resuspend the cells in SC-specific media.
Transfer the cells to an uncoated plate and incubate.
Any remaining fibroblasts will adhere, leaving SCs in suspension.
Transfer the suspension to a matrix-coated plate.
Incubate for Schwann cell attachment and division.
