NPC Neurosphere Assay: An In Vitro Assay to Assess the Migratory Response of Neural Precursor Cells from the Neurosphere
Published: April 30, 2023
Abstract
Source: Williams, M. et al. Rapid Detection of Neurodevelopmental Phenotypes in Human Neural Precursor Cells (NPCs). J. Vis. Exp. (2018)
This video demonstrates the neural precursor cell neurosphere migration assay—an in vitro assay to study the migratory response of neural precursor cells from the neurosphere—via microscopic analysis.
Protocol
1. NPC Neurosphere Migration Assay Neurosphere Formation Add 1 mL of 100% Expansion Media into a 35 mm dish with no coating substrate. Incubate dishes for at least 15 min at 37 °C before plating NPCs. Prepare 2–3 dishes to ensure that there will be enough neurospheres for the cell migration assay. NOTE: The absence of a coating substrate ensures that NPCs remain suspended in the media, which is essential for neurosphere formatio…
Representative Results
Figure 1: Selecting neurospheres for migration assay. (A) Phase-contrast image of a representative field of neurospheres at 24 h. Spheres are all less than 100 μm and thus, are not collected for the migration assay. (B) Phase-contrast image of a representative field of neurospheres at 72 h. All spheres are within the 100 μm ± 20 µm range, indicating they are ready to be collected…
Açıklamalar
The authors have nothing to disclose.
Materials
| Neurobasal Medium | ThermoFischer Scientific | 21103049 | Component of both NIM and 100% Expansion Medium |
| PSC Neural Induction Medium: Protocol Link: https://goo.gl/ euub7a | ThermoFischer Scientific | A1647801 | This is a kit that consists of Neurobasal (NB) medium and a 50x Neural Induction Supplement (NIS). The NIS is used to make 1X Neural Induction Medium and 100% Expansion Medium |
| Advanced DMEM/F12 Medium | ThermoFischer Scientific | 12634-010 | Component of 100% Expansion Medium |
| hESC-qualified Matrigel | Corning | 354277 | hESC-qualified extracellular matrixmimic gel (ECM-mimic gel) |
| 6-well plates | Corning | COR-3506 | Polystyrene plates used for NPC maintenance and for Neurosphere Migration Assay |
| Human Basic FGF-2 | Peprotech | 100-18B | Growth factor |
| Penicillin/Streptomycin | ThermoFischer Scientific | 15140122 | Antibiotic, component of NIM, 100% Expansion and 30% Expansion Media |
Etiketler
Bu Makaleden Alıntı Yapın
NPC Neurosphere Assay: An In Vitro Assay to Assess the Migratory Response of Neural Precursor Cells from the Neurosphere. J. Vis. Exp. (Pending Publication), e20630, doi: (2023).