Improvement of cellular pattern organization and clarity through centrifugal force

Rapid and strategic cell placement is necessary for high throughput tissue fabrication. Current adhesive cell patterning systems rely on fluidic shear flow to remove cells outside of the patterned regions, but limitations in washing complexity and uniformity prevent adhesive patterns from being widely applied. Centrifugation is commonly used to study the adhesive strength of cells to various substrates; however, the approach has not been applied to selective cell adhesion systems to create highly organized cell patterns. This study shows centrifugation as a promising method to wash cellular patterns after selective binding of cells to the surface has taken place. After patterning H9C2 cells using biotin-streptavidin as a model adhesive patterning system and washing with centrifugation, there is a significant number of cells removed outside of the patterned areas of the substrate compared to the initial seeding, while there is not a significant number removed from the desired patterned areas. This method is effective in patterning multiple size and linear structures from line widths of 50-200 μm without compromising immediate cell viability below 80%. We also test this procedure on a variety of tube-forming cell lines (MPCs, HUVECs) on various tissue-like surface materials (collagen 1 and Matrigel) with no significant differences in their respective tube formation metrics when the cells were seeded directly on their unconjugated surface versus patterned and washed through centrifugation. This result demonstrates that our patterning and centrifugation system can be adapted to a variety of cell types and substrates to create patterns tailored to many biological applications.