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Material surface engineering for multiplex cell culture in microwell

Recent developments in cell-based assays and high throughput screening include living-cell chips, allowing the delivery of experimental stimuli and measurement of the resulting cellular responses. We have been developing a new concept for localized cells co-culture. This cell-chip consists of a polystyrene spin-coated solid support bearing gold bottomed microwells. The cell chip support is fabricated as follows: (i) electro-sputtering of a thin layer of gold (40 nm) onto a polycarbonate substrate, (ii) spin-coating of a polystyrene thin film (500 nm ± 50 nm) over the gold layer, followed by (iii) polystyrene etching through the spotting of toluene nanovolume (300-900 pL). In each gold bottomed microwell, a small population of adherent cells (approx. 100 cells) can be cultured. In this miniaturized system, different cell lines can be co-cultured on a 1 cm² surface, opening the way to multiplexed cell-chip development. In order to keep the cells in a properly hydrated environment and to physically retain them before they adhere, a biocompatible alginate polymer was used during the robotized micro pipetting. This approach allows the encapsulation of the cell in a very small volume (50 nL), directly in the microwells. After 24 hours of culture, the cells adhered on the gold bottom of the microwells and the alginate matrix was removed by addition of calcium-free culture medium.