Isolation of Stem Cells from Postnatal Mouse Cerebellum and Differentiation into Neural Cells
Trascrizione
Take cells isolated from a newborn mouse cerebellum containing stem cells, neurons, and glia.
Add antibody-coated magnetic microbeads that bind to a stem-cell-specific surface glycoprotein.
Centrifuge, remove the unbound beads, and resuspend the cells in a buffer.
Load the cells onto a separation column placed in a magnetic separator, which retains the microbead-bound stem cells while other cells flow through.
Add a neurosphere medium to elute the cells and transfer them into a microplate. Incubate to induce the formation of primary neurospheres, self-renewing colonies of stem cells.
Centrifuge and discard the medium. Resuspend the neurospheres in a medium containing enzymes to digest the cellular adhesion proteins.
Centrifuge and remove the enzymes. Add the neurosphere medium and mechanically dissociate the cells.
Incubate to allow stem cell proliferation and secondary neurosphere formation, while the differentiated cells lacking stem cell characteristics do not proliferate.
Transfer the neurospheres to a differentiation medium and incubate, promoting differentiation into neurons and glia.
