Analysis of invasive activity of CAF spheroids into three dimensional (3D) collagen matrices

María Ángeles Villaronga*, Saúl Álvarez Teijeiro, Francisco Hermida-Prado, Marta Garzón-Arango, Victoria Sanz-Moreno, Juana María García-Pedrero

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

5 Citations (Scopus)


Tumor growth and progression is the result of a complex process controlled not only by malignant cancer cells but also by the surrounding tumor microenvironment (TME). Cancer associated fibroblasts (CAFs), the most abundant cellular component of TME, play an active role in tumor invasion and metastasis by promoting cancer cell invasion through cell-cell interactions and secretion of pro-invasive factors such as extracellular matrix (ECM)-degrading proteases. Due to their tumor-promoting activities, there is an emerging interest in investigating CAFs biology and its potential as drug targets for cancer therapies. Here we describe an easy and highly reproducible quantitative method to analyze CAF invasive activity by forming multicellular spheroids embedded into a three-dimensional (3D) matrix that mimics in vivo ECM. Subsequently, invasion is monitored over time using a time-lapse microscope. We also provide an automated image analysis system that enables the rapid quantification of the spheroid area increase (invasive area) over time. The use of a 96-well plate format with one CAF spheroid per well and the automated analysis provides a method suitable for drug screening test, such as protease inhibitors.

Original languageEnglish
Title of host publicationProteases and Cancer: Methods and Protocols
EditorsSantiago Cal, Alvaro J. Obaya
PublisherHumana Press Inc
Number of pages10
Publication statusE-pub ahead of print - 10 Jan 2018

Publication series

NameMethods in Molecular Biology
ISSN (Print)1064-3745


  • Cancer associated fibroblasts
  • Collagen
  • Extracellular matrix
  • Invasive activity
  • Proteases
  • Spheroids


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