A novel three-dimensional cell culture system was constructed with an array of cell panels (4 x 5) in a silicon chip, together with multi-channel drug containers. Human breast cancer (MCF-7) cells were embedded in a collagen-gel matrix and entrapped in a pyramidal-shaped silicon hole. Each cell panel can be isolated by a channel composed of a microfluid part and a reservoir. A cell panel was exposed to 200 mm KCN for 2 days to demonstrate that each cell panel could be independently evaluated under various stimulation conditions. Based on the cellular respiration activity, the proliferation behavior was continuously monitored on the silicon-based cell array for 5 days using scanning electrochemical microscopy (SECM). The cells entrapped in the device (3-D culture) proliferated normally, and the proliferation rate was lower than that of cells grown in a monolayer cell culture (2-D culture). The effects of three anticancer drugs measured simultaneously on the cell chip were in good agreement with those obtained by a conventional colorimetric assay. Our results suggest that the silicon-based device for 3D culture is appropriate for a chemosensitivity assay involving multi-chemical stimulation.