Imaging- and Flow Cytometry-based Analysis of Cell Position and the Cell Cycle in 3D Melanoma Spheroids

Journal of Visualized Experiments : JoVE
Kimberley A BeaumontNikolas K Haass

Abstract

Three-dimensional (3D) tumor spheroids are utilized in cancer research as a more accurate model of the in vivo tumor microenvironment, compared to traditional two-dimensional (2D) cell culture. The spheroid model is able to mimic the effects of cell-cell interaction, hypoxia and nutrient deprivation, and drug penetration. One characteristic of this model is the development of a necrotic core, surrounded by a ring of G1 arrested cells, with proliferating cells on the outer layers of the spheroid. Of interest in the cancer field is how different regions of the spheroid respond to drug therapies as well as genetic or environmental manipulation. We describe here the use of the fluorescence ubiquitination cell cycle indicator (FUCCI) system along with cytometry and image analysis using commercial software to characterize the cell cycle status of cells with respect to their position inside melanoma spheroids. These methods may be used to track changes in cell cycle status, gene/protein expression or cell viability in different sub-regions of tumor spheroids over time and under different conditions.

Citations

Mar 13, 2016·The Journal of Investigative Dermatology·Kimberley A BeaumontNikolas K Haass
Apr 26, 2019·Cell Death & Disease·Erinna F LeeW Douglas Fairlie
Dec 10, 2020·Pharmaceutics·Bárbara PintoHassan Bousbaa
Mar 20, 2021·Proceedings of the National Academy of Sciences of the United States of America·Stefania IppatiLars M Ittner

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