To directly observe transendothelial migration of cancer cells, we performed a microfluidic‐based 3D tumor assay for HDLEC intravasation that enabled the real‐time visualization and quantification of the interactions between cancer cells and HDLEC monolayer that were stimulated with 50 ng·mL−1 periostin for 2 days. This evidence concerns the gene POSTN and neoplasm.