Brain Tumor Stem Cell (BTSC) Neurosphere Invasion Assay: An In Vitro Assay to Analyze the Effect of Drug Treatment on BTSC Invasion from Neurosphere

Abstract

Source: Restall, I. et al. Live-Cell imaging assays to study glioblastoma brain tumor stem cell migration and invasion. J. Vis. Exp. (2018)

This video demonstrates the in vitro brain tumor stem cell (BTSC) invasion assay to determine the effect of drug treatment on the invasion capacity of BTSCs from the neurosphere via live-cell imaging technique.

Protocol

1. Brain Tumor Stem Cell Neurosphere Invasion Assay

1. Seed 200,000 cells in 8 mL of complete media in a T25 flask 1–2 weeks before planning to perform an invasion assay.
   NOTE: The timing from plating to performing the assay will depend on the proliferation rate of the BTSC culture used.
   1. For pretreated cells, add the compound of interest to the flask at the desired concentration for the predetermined time course before the neurospheres are ready to be plated for invasion.
2. Wait until the average neurosphere size is approximately 150–200 μm; typically, this takes 7R....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Rat Collagen I	Trevigen	3440-100-01
NaOH	VWR	BDH3247-1
Acetic Acid	Millipore Sigma	AX0073-6
96-well plates	Eppendorf	30730119
T25 Nunc EasYFlask	ThermoFisher	156367
Falcon 15 mL conical tube	ThermoFisher	352096
Incucyte ClearView 96-Well Chemotaxis plate	Essen Bioscience	4582
IncuCyte Zoom Live-Cell Analysis System	Essen Bioscience	4545
IncuCyte Zoom Live-Cell Analysis Software	Essen Bioscience	2016A Rev1
Penicillin, streptomycin	Sigma	P4333
Phosphate buffered saline (PBS)	Sigma	D8537
Fetal bovine serum (FBS)	Invitrogen	12483-020