University of Oulu

Naakka, E., Tuomainen, K., Wistrand, H., Palkama, M., Suleymanova, I., Al-Samadi, A., Salo, T. Fully Human Tumor-based Matrix in Three-dimensional Spheroid Invasion Assay. J. Vis. Exp. (147), e59567, doi:10.3791/59567 (2019).

Fully human tumor-based matrix in three-dimensional spheroid invasion assay

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Author: Naakka, Erika1; Tuomainen, Katja1; Wistrand, Henrik1;
Organizations: 1Department of Oral and Maxillofacial Diseases, Clinicum, Faculty of Medicine, University of Helsinki
2Cancer and Translational Medicine Research Unit, University of Oulu
3Medical Research Center, Oulu University Hospital
4Helsinki University Hospital
Format: article
Version: published version
Access: open
Online Access: PDF Full Text (PDF, 5.5 MB)
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Language: English
Published: MyJoVE Corporation, 2019
Publish Date: 2019-06-14


Two-dimensional cell culture-based assays are commonly used in in vitro cancer research. However, they lack several basic elements that form the tumor microenvironment. To obtain more reliable in vitro results, several three-dimensional (3D) cell culture assays have been introduced. These assays allow cancer cells to interact with the extracellular matrix. This interaction affects cell behavior, such as proliferation and invasion, as well as cell morphology. Additionally, this interaction could induce or suppress the expression of several pro- and anti-tumorigenic molecules. Spheroid invasion assay was developed to provide a suitable 3D in vitro method to study cancer cell invasion. Currently, animal-derived matrices, such as mouse sarcoma-derived matrix (MSDM) and rat tail type I collagen, are mainly used in the spheroid invasion assays. Taking into consideration the differences between the human tumor microenvironment and animal-derived matrices, a human myoma-derived matrix (HMDM) was developed from benign uterus leiomyoma tissue. It has been shown that HMDM induces migration and invasion of carcinoma cells better than MSDM. This protocol provided a simple, reproducible, and reliable 3D human tumor-based spheroid invasion assay using the HMDM/fibrin matrix. It also includes detailed instructions on imaging and analysis. The spheroids grow in a U-shaped ultra-low attachment plate within the HMDM/fibrin matrix and invade through it. The invasion is daily imaged, measured, and analyzed using ilastik and Fiji ImageJ software. The assay platform was demonstrated using human laryngeal primary and metastatic squamous cell carcinoma cell lines. However, the protocol is suitable also for other solid cancer cell lines.

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Series: Journal of visualized experiments
ISSN: 1940-087X
ISSN-E: 1940-087X
ISSN-L: 1940-087X
Volume: 147
Article number: e59567
DOI: 10.3791/59567
Type of Publication: A1 Journal article – refereed
Field of Science: 3122 Cancers
Funding: The authors thank the funders of this study: the Sigrid Jusélius Foundation, the Cancer Society of Finland, and Helsinki University Central Hospital research funds. The authors acknowledge the Biomedicum Imaging Unit at the University of Helsinki for technical help.
Copyright information: © 2019 Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported Licens