Live Imaging of Cortical Neural Stem Cell Neurite Outgrowth with IncuCyte NeuroTrack
1) Differentiation and quantification:
- To perform a morphological characterization of Axol Cortical Neural Stem Cells throughout the differentiation process and understand the suitability of Essen's phase contrast NeuroTrackTM analysis algorithm.
- Method development (cell density, coating conditions, cell feeding protocol).
2) Passaging post-differentiated cells:
- To assess the possibility of passaging the cells post plating to create a model to study both neurite outgrowth as well as neurodegeneration.
3) Cell Type Comparison:
- To complete a time-course comparison of neurite outgrowth using N2a cells, Primary cells, Competitor iPSC-derived neurons and Axol Cortical Neural Stem Cells-derived Neurons.
Differentiation & Quantification
Differentiation of Axol Cortical Neural Stem Cells
Axol cortical NSCs were successfully differentiated into human cerebral cortical neurons following treatment with Axol Neural AdvanceTM.
Passaging of Post-Differentiated Cells
- Cells were defrosted into 35-mm dish and differentiated for 4 days as per protocol.
- Cells were lifted with Axol UnlockTM and seeded into 1x 96-well TPP plate in Axol Sure BoostTM.
- 24 h post-plating 100 % of the media was replaced.
Neurites establishment occurred within 6 h post-plating.
Cell Type Comparison
- Time-courses compare the mean ± SEM of NL, BP and NL/CBC values for rat primary cortical neurons, competitor iPSC-derived neurons, Axol-hNPC-derived neurons and N2a cells.
- High levels of neurites can be quantified using Zoom algorithm (60 - 70 mm/mm2, compared to 25 – 30 mm/mm2 for competitor iPSC-derived neurons).
- Overall, Axol NSCs yielded high Neurite length and branch point values when compared to any other cell type.
- Differentiated Axol NSCs can be passaged with a high survival rate.
- Onset of neurite outgrowth occurs within 6 h post passaging.
- NSCs yielded high Neurite length and branch point values when compared to competitor iPSC-derived neurons, primary rat cortical neurons or N2a cells (~ 60, ~25, ~30, ~20 mm/mm2, respectively).