Transplantation of Stem Cell-Derived Neuronal Progenitors into Human Cortical Slices

Begin with an adherent culture of stem cell-derived neuronal progenitor cells primed for differentiation into cortical neurons.

Remove the medium, then add an enzyme to disrupt the extracellular matrix and detach the cells.

Use an inhibitor to neutralize the enzyme, then add a fresh medium.

Transfer the suspension into a tube, centrifuge, and discard the supernatant to isolate the cells.

Resuspend the cells in a basement membrane matrix to facilitate transplantation into brain tissue, then load them into a cold capillary tube.

Take a human cortical slice onto a transwell assembly containing a medium.

Remove part of the medium, then inject droplets of the cell suspension into the tissue.

Incubate to allow the matrix to solidify.

Add a medium to immerse the tissue, then incubate to enable progenitor cells to differentiate into neurons, extend their neurites, and form synapses with host cortical neurons.

The cortical slice is now ready for analysis.

Remove the media from the cell culture flask. On day seven of differentiation, detach cortically-primed GFP-lt-NES cells by adding 500 microliters of trypsin and incubate for 5 to 10 minutes at room temperature.

Next, add an equal volume of the tryps