Differentiating Human Neural Stem Cells into Neural and Astrocyte Progenitors
Transcription
Begin with non-adherent neurospheres, a three-dimensional aggregate of human neural stem cells in a cell culture-derived conditioned medium.
This medium contains nutrients, growth factors, and extracellular matrix components that maintain cell viability.
Centrifuge to collect the neurospheres.
Treat them with trypsin and DNase enzymes, mix the contents, and then incubate.
Trypsin disrupts cell-to-cell adhesion and facilitates cell detachment, while DNase degrades free DNA to prevent cell clumping.
Pipette the contents repeatedly to form a single-cell suspension. Then, add a trypsin inhibitor to neutralize the enzyme activity.
Centrifuge and remove the supernatant, and then resuspend the cells in a proliferation medium containing neural growth factors.
Seed the cell suspension onto a polymer-coated coverslip placed in a multi-well plate. Incubate to allow the stem cell attachment to the polymer.
Further, based on the absorption of specific growth factors, the stem cells differentiate into neural and astrocyte progenitors, which are ready for further maturation into specialized cell types.
