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To investigate tumor cell migration capability, the scratch/wound healing assay and the Transwell assay are the most commonly used assays in the current biomedical research laboratory. However, both assays have their limitations and may mislead the interpretation of the results. In the current study, visualization and quantification of tumor cell migration process was realized in a three-dimensional (3D) environment. The tumor cells horizontally migrated along a Matrigel-filled microchannel under extracellular stimulation. The cell migration process was visualized under a microscope, and the migration speed could be calculated based on the traveling distance of the cells and the time required. Here, three demonstrations were conducted, respectively, including cells attracted by nutrient gradient, stimulated by cytokine, and coculturing with fibroblasts. The results revealed that the cell migration capability could be visually and quantitatively correlated to the extracellular stimulation. The current protocol is compatible to the existing laboratory setup and provides a persuasive result for the study of the 3D cell migration process. Understanding of the molecular and intercellular mechanism of cancer metastasis can potentially develop effective therapeutic strategy.

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