Differentiation of a Human Neuroblastoma Cell Line into Mature Neurons
Transcription
Begin with an undifferentiated neuroblastoma cell line comprising neuron-like and epithelial-like cells in a high-serum-concentration medium.
Remove the medium and add a differentiation medium containing a reduced serum concentration, supplemented with retinoic acid.
Serum deprivation prevents epithelial-like cell proliferation.
Retinoic acid induces neuronal gene expression in the neuron-like cells, transforming them into a more differentiated phenotype.
Add a trypsin-EDTA cocktail to detach the partially differentiated cells.
Add media containing serum proteins to block trypsin.
Plate the cells and introduce a fresh differentiation medium containing retinoic acid and a further reduced serum level.
The cells continue to develop extended processes.
Add trypsin to detach the cells.
Next, add media to block trypsin and plate the cells on an extracellular matrix protein-coated dish to promote cellular adhesion.
Finally, supplement with serum-free medium containing neuronal growth factors.
These factors induce terminal differentiation into mature neurons exhibiting extensive branched neuritic processes.
