Heterotypic cell pair co-culturing on patterned microarrays

Abstract

We present a pair-wise co-culturing technique that creates large numbers of heterotypic cell pairs in patterned arrays. Lithographic patterning produces arrays with thousands of traps, each designed to accommodate only two cells and confine them at these sites for co-culturing. Two variants are introduced: a random seeding method that sediments a mixture of two cell types onto the array, and an approach that incorporates ferromagnetic thin films into the arrays and attracts cells that have been attached to ferromagnetic nanowires to the array sites through dipole interactions. The array technique includes the utilization of custom image analysis software that extracts data from multi-channel fluorescence images and records information about the cells in every trap, enabling the acquisition of accurate, high-statistics data. The applicability of the technique was demonstrated in experiments examining proliferation rates in pairs of bovine pulmonary artery endothelial and smooth muscle cells. Results demonstrated that heterotypic interactions favored smooth muscle cell proliferation while disfavoring endothelial cell proliferation. This is one example of a variety of cell-cell interactions that could be probed with this method.

Co-culture systems and technologies: taking synthetic biology to the next level

“…Co-cultures have also been performed on solid media, such as Petri dishes [10,28,34,40,48,83], slides [71,77,78,81,82] or within cell patterning chambers encased in hydrogel [57]. Asymmetric culture volumes for the two populations are also possible [80,87].…”

“…Examples of co-cultures of more diverse populations include cultures of cells from different species of multicellular eukaryotic organisms [77][78][79][80][81][82][83], fungi [84] or different bacterial species [10,18,22,34,55,85,86]. Mammalian and bacterial cells have been co-cultured, either for infection studies [1,87,88] or testing of synthetic bacteria [35,36,89].…”

“…For co-culturing populations with a degree of separation, commonly used options include transwell plates [56,103], microfluidic platforms [4,6,94,96] or solid supports such as Petri dishes [10,40], three-dimensional scaffolds [74], hydrogels [57] or microarrays [82]. Many bespoke microfluidic rsif.royalsocietypublishing.org J. R. Soc.…”

“…In other studies, contact between cells is allowed, but carefully controlled through techniques, such as cell immobilization [82], microfluidics [6,58,73] or cell micropatterning [54,71,72,77,78,81,92] (figure 3). The type of micropattern used can strongly affect the culture fate [72,77], because it will determine diffusion rates and level of separation or contact between cell populations.…”

“…(c) Co-cultures on solid supports. Using gels [57], droplets [68], microarrays [82] or cell layers on glass slides [81]. (d ) Co-culture systems using bioreactors, with possibility for separation between population compartments [17,67,70].…”