Combination of 3D Micro Contact Printing and Scraping Technique for Site Selective Cocultivation Across a Porous Micro Structured Scaffold

3D-microcontact printing (3D-µCP) technique combines the advantages of microcontact printing and microthermoforming for the fabrication of functional biomaterials with complexity closer to real tissue. Despite its unmet advantages in terms of complexity and processability in a single step, this technique is limited to the front side of the substrate. Considering the advantage of inherent topography patterns on the reverse side of the substrate, an additional degree of patterning can be envisioned. However, selective patterning on the reverse side is challenging due to the fragility of the cell culture on the front side. Herein, a technically simple scraping technique is presented in combination with 3D-µCP to generate 3D cell patterns on both the front and the reverse side of a micro thermoformed substrate. The technical advancement of 3D-µCP with the scraping technique offers a complex, multi-layered co-culture system that can be used to study cell-to-cell crosstalk at the microlevel with topographies similar to real tissue. Direct seeding of the third cell type on the scaffold expands cell type complexity, resulting in a tri-culture model. As a proof of concept, a triculture of EA.Hy 926 endothelial cells, HepG2 hepatoblastoma cells, and NIH3T3 fibroblasts are generated, mimicking the tumor microenvironment in the liver.