Spherical Invasion Assay: A novel method to measure the pro‐invasive activity of cancer cells

Abstract

Tumor invasion refers to the process by which cancer cells penetrate the basement membrane into surrounding stroma, neighboring blood vessels and lymph nodes. Tumor invasion is an essential step for metastasis of human cancers. The Boyden Chamber assay is the benchmark technique of measuring the invasion of cancer cells. However, this assay suffers from several caveats. The physiological relevance of transwell inserts (to recapitulate invasion in vivo) is poor. The pore size of the membrane highly influences the number of invaded cells. Another limitation of the Boyden chamber assay is it is very difficult to control the concentration of the chemokine gradient and this may produce aberrant results. All these considerations led us to develop a novel method of measuring tumor invasion namely the SPHERICAL INVASION ASSAY (SIA) in our laboratory. The SIA measures the invasion of human cancer cells as they migrate from the primary Matrigel layer, over the interface and travel into a secondary Matrigel layer. The cells are grown in extracellular matrix (ECM) retain biological characteristics of tumors, such as diffusion gradient of oxygen, nutrients and pH. The growth of the cells inside the ECM allows for complex cell‐cell and cell‐matrix interaction. Most importantly, the results of the SIA correlate with the data obtained in the Boyden Chamber assay, so the SIA can be used to confirm the results obtained in Boyden Chamber assays1. The present protocol examines the effect of the Src Kinase inhibitor PP2 on the invasion of A549 human NSCLC cells. We hope that the SIA will be useful tool for researchers working in the field of tumor microenvironment biology and cancer metastasis.