Polymeric Aqueous Biphasic Systems for Non‐Contact Cell Printing on Cells: Engineering Heterocellular Embryonic Stem Cell Niches

Abstract

An optimized polymeric aqueous two‐phase system allows direct and non‐contact printing of cells onto a monolayer of living cells in arbitrary shapes as well as in a high‐density microarray format to create heterocellular microenvironments and study the effect of direct cell–cell interactions on cell fate. The entire process is performed in aqueous media to support full cell viability and functionality.

Cell Co-culture Patterning Using Aqueous Two-phase Systems

“…This method allows any researcher with access to a typical cell culture lab (access to a hood, CO 2 incubator, and micropipettes) and the aforementioned polymers to reproducibly pattern cells in monoculture and co-culture. Our lab has demonstrated this capability by printing arrays of cells to study cell migration in a wound healing assay and to examine the effects of juxtacrine and paracrine signaling in the differentiation of embryonic cells 16,17,20 . Other methods, including patterning of extracellular matrix 28 , inkjet printing 29 , and patterning by laminar flow in microfluidic devices 25,26 have also been used to localize cells.…”

“…When cells are incorporated into the PEG phase, they are excluded from entering the DEX droplets due to PEG/DEX interfacial tension 20 . When cells are patterned in the DEX phase, they are retained at the surface of the cell culture substrate by interfacial tension and partitioning 16,17,19 .…”

“…ATPSs formed by PEG and DEX generally display interfacial tensions that are much lower than other liquid-liquid two-phase systems such as oil and water; however, the interfacial tension forces still exert effects on small particles such as viruses, cells and protein aggregates 2,[11][12][13] . Finally, since higher molecular weight PEG and DEX separate at low concentrations (less than 5% wt/wt for high molecular weight polymers varieties) in the presence of physiological concentrations of salts, there are few if any deleterious effects on mammalian cells incorporated within these systems [14][15][16] . Recently, the interfacial properties and partitioning effects of ATPSs have been applied by our lab for cell patterning 14,[16][17][18][19][20] .…”

“…Finally, since higher molecular weight PEG and DEX separate at low concentrations (less than 5% wt/wt for high molecular weight polymers varieties) in the presence of physiological concentrations of salts, there are few if any deleterious effects on mammalian cells incorporated within these systems [14][15][16] . Recently, the interfacial properties and partitioning effects of ATPSs have been applied by our lab for cell patterning 14,[16][17][18][19][20] . This was accomplished by micropatterning a denser DEX solution on cell culture substrates in the presence of PEG.…”

An open-chamber flow-focusing device for focal stimulation of micropatterned cells

“…Therefore, microfluidic devices provide key advantages over traditional cell culture platforms in their ability to recapitulate the physiological conditions at the microscale. 2,3,6,7 Microfluidic manipulation has been utilized for the focal stimulation of microdomains of single cells, subcellular compartments, or tissues in a wide range of applications including: in vitro models for tissue development that necessitate spatiotemporal presentation of soluble and physical signals, 8,9 single cell analysis for multiplexed drug testing, 10,11 localized chemical stimulation of tissue slices for neural electrophysiology and cancer drug screening, [12][13][14] and localized neurochemical stimulation of muscle cells. [15][16][17] Classical microfluidic approaches for localized stimulation in cellular assays have utilized pressure-driven laminar flow in enclosed channels.…”