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The interaction of cancer cells with the stromal cells and matrix in the tumor microenvironment plays a key role in progression to metastasis. A better understanding of the mechanisms underlying these interactions would aid in developing new therapeutic approaches to inhibit this progression. Here, we describe the fabrication of a simple microfluidic bioreactor capable of recapitulating the three-dimensional breast tumor microenvironment. Cancer cell spheroids, fibroblasts, and endothelial cells co-cultured in this device create a robust microenvironment suitable for studying in real time the migration of cancer cells along matrix structures laid down by fibroblasts within the 3D tumor microenvironment. This system allows for ready evaluation of response to targeted therapy.