Isolation and Culturing of Adult Neural Stem Cells from the Mouse Subcallosal Zone

Begin with a mouse brain on a chilled brain matrix.

Obtain sections containing the subcallosal zone or SCZ. Transfer the section to a cold phosphate buffer.

Under a dissecting microscope, cut out the SCZ portion.

Chop the dissected SCZ into smaller pieces.

Transfer these pieces into a digestion buffer. Incubate to dissociate the cells from the tissue.

Centrifuge and discard the supernatant. Resuspend the cells in a suitable medium.

Transfer the cells to a culture plate and add specific growth factors.

The adult neural stem cells divide, forming clusters called neurospheres.

Improper nutrient distribution in cell clusters limits cell growth.

To disperse the clusters, centrifuge and discard the supernatant. Resuspend the clusters in a dissociation buffer and incubate.

Pipette the solution up and down to generate single cells.

Centrifuge and discard the supernatant.

Resuspend the cells and transfer them into a suitable medium to obtain a monolayer.

Once dissected, transfer the mouse brain to cold PBS buffer, and rinse it twice to remove excess blood. Next, place the brain in the brain matrix on ice, and make coronal cuts to obtain one-millimeter-thick slices. Subsequently, transfer th