Neural Progenitor Cells Derived from ATCC-DYS0530 Parkinson's Disease
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Cells contain Sendai virus (SeV) DNA sequences
ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.
Complete dopaminergic differentiation medium for NPCs is DMEM: F12 (ATCC 30-2006); supplemented with the Neural Progenitor Cell Dopaminergic Differentiation Kit (ATCC ACS-3004). To make complete NPC dopaminergic differentiation medium add the following components of this kit to 237 mL DMEM: F12:
Coat plates with Cell Basement Membrane Gel (ATCC ACS-3035) and culture the NPCs with NPC Growth Medium (ATCC ACS-3003) to provide a surface for the attachment of NPCs.
Post thaw day 1, perform a 100% medium change and remove all cells that did not attach. Perform a 100% medium change every other day thereafter. Passage the cells cells with diluted Accutase (50% Accutase and 50% DPBS) when they reach ~95% confluence and reseed the NPCs at 40,000 viable cells/cm2 on Cell Basement Membrane-coated dishes/plates.
Dopaminergic Differentiation: Seed NPCs at 10,000 cells/cm2 in a 12-well plate pre-coated with Cell Basement Membrane. Culture overnight in complete NPC expansion medium. The next day, aspirate the medium and add 1.5 ml of pre-warmed complete Dopaminergic Differentiation Medium containing CHIR. Change the Dopaminergic media every other day (e.g., Monday, Wednesday, and Friday) for 3 weeks as the following:
Astrocyte, oligodendrocyte, and neuron differentiation; drug screening
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Malik N, et al. Compounds with species and cell type specific toxicity identified in a 2000 compound drug screen of neural stem cells and rat mixed cortical neurons. Neurotoxicology 45: 192-200, 2014. PubMed: 25454721