Differentiating Mouse Embryonic Stem Cells into a Networked Neuron Population
Transcrição
Seed mouse embryonic stem cells in a low-attachment dish containing differentiation media.
Incubate under rotation to facilitate cellular aggregation and differentiation into a neuronal lineage.
Transfer the aggregates to a tube and allow them to settle by gravity. Discard the media.
Resuspend and transfer the aggregates to the original dish containing differentiation media supplemented with retinoic acid.
Retinoic acid induces neuronal gene expression, triggering differentiation into neural progenitor cells.
Transfer the aggregates to a tube. Remove the media and add trypsin to dissociate the aggregates.
Add an inhibitor to neutralize the trypsin.
Pipette repeatedly to obtain a homogeneous cell suspension. Filter the suspension to remove aggregates.
Centrifuge and remove the supernatant. Resuspend the cells in media.
Seed the cells on a polymer-coated dish to facilitate cell adhesion.
Remove the media. Add fresh media containing growth factors to promote neuronal maturation and form a networked neuron population.
