Luminescence-Based Assay to Assess Neurotoxicity of Test Compounds on Neurons

Published: April 30, 2023

Abstract

Source: Wahlestedt, C., et al., A Neurite Outgrowth Assay and Neurotoxicity Assessment with Human Neural Progenitor Cell-Derived Neurons. J. Vis. Exp. (2020)

In this video, we describe a luminescence-based method to assess the neurotoxicity of neurite outgrowth-promoting small molecule compounds on the viability of hNPC-derived neurons. The assay measures intracellular ATP level, an indicator of cell viability, to evaluate neurotoxicity following test compound treatment.

Protocol

All procedures involving human participants have been performed in compliance with the institutional, national, and international guidelines for human welfare and have been reviewed by the local institutional review board 1. Isolation and culture of human neural progenitor cells (hNPCs) Place the brain tissue in a 100 mm Petri dish and carefully remove the meninges using forceps. Transfer the brain tissue to a 50 mL conical tube and wash it twice with 20 mL of PBS by gent…

Offenlegungen

The authors have nothing to disclose.

Materials

Antibiotic-Antimycotic Gibco 15240062
B27 Thermo 17504001
B27 – minus vitamin A Thermo 12587010
BDNF PeproTech 450-02
BSA Sigma A8531
CellTiter-Glo Promega G7572
CoolCell Corning 432000 Cell freezing containers ensuring standardized controlled-rate -1°C/minute cell freezing in a -80°C freezer
CryoStor CS10 StemCell Technologies 7930 Cryopreservation medium containing 10% DMSO
DMEM/F12 Gibco 11320033
DMSO Sigma 34869-100ML
EGF Gibco PHG0311
FGF Gibco PHG6015
GDNF PeproTech 450-10
Glutamax Gibco 35050061 L-alanyl-L-glutamine supplement
Heparin Calbiochem 375095
Laminin Sigma L2020-1MG
L-Ascorbic Acid Sigma A92902-25G
mFreSR StemCell Technologies 5854 Serum-free cryopreservation medium designed for the cryopreservation of human embryonic and induced pluripotent stem cells
N2 Gibco 17502048
Nunc 384-Well Polystyrene White Microplates Thermo 164610
PBS Thermo 10010-049
Poly-L-lysine Sigma P5899-5MG
StemPro Accutase Gibco A1110501 Cell dissociation reagent containing proteolytic and collagenolytic enzymes
Neurobasal Medium Gibco 21103049

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Luminescence-Based Assay to Assess Neurotoxicity of Test Compounds on Neurons. J. Vis. Exp. (Pending Publication), e21330, doi: (2023).

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