Controlled Synthesis of Cell-Laden Microgels by Radical-Free Gelation in Droplet Microfluidics

Abstract

Micrometer-sized hydrogel particles that contain living cells can be fabricated with exquisite control through the use of dropletbased microfluidics and bioinert polymers such as polyethyleneglycol (PEG) and hyperbranched polyglycerol (hPG). However, in existing techniques, the microgel gelation is often achieved through harmful reactions with free radicals. This is detrimental for the viability of the encapsulated cells. To overcome this limitation, we present a technique that combines droplet microfluidic templating with bio-orthogonal thiol−ene click reactions to fabricate monodisperse, cell-laden microgel particles. The gelation of these microgels is achieved via the nucleophilic Michael addition of dithiolated PEG macro-cross-linkers to acrylated hPG building blocks and does not require any initiator. We systematically vary the microgel properties through the use of PEG linkers with different molecular weights along with different concentrations of macromonomers to investigate the influence of these parameters on the viability and proliferation of encapsulated yeast cells. We also demonstrate the encapsulation of mammalian cells including fibroblasts and lymphoblasts.

Microfluidic Synthesis of Cell-Type-Specific Artificial Extracellular Matrix Hydrogels

“…42−44 As a result, the size distribution of microgels that was obtained after removal of the oil phase was very narrow ( Figure 2B). 35,44 The chosen channel geometry is particularly well suited for the generation of multicomponent microgels that can, for example, be obtained by loading different gel components into the precursors streams. For instance, in order to facilitate medium exchange during bioreactor cell culture, magnetic particles were loaded into one of the precursor streams to irreversibly trap them in the microgels.…”

Poly(vinyl alcohol)-heparin biosynthetic microspheres produced by microfluidics and ultraviolet photopolymerisation

“…Microfluidics has previously been used with alginate to encapsulate cells with high viability [23][24][25] and more recently with thiol-ene click reactions with PEG. 17 UV polymerisation is a common method for encapsulating cells in bulk gels and has been used with many cell types including chondrocytes 49 and mesenchymal stem cells 50 with high viability. Additionally, L929 fibroblasts that were encapsulated in the same PVA microspheres produced by a UV photopolymerisation and electrospray method also had >90% viability at 24 h. 8 Heparin has been incorporated into bulk hydrogels for the purpose of sustained growth factor release and presentation to cells and was shown to have beneficial effects on viability.…”

“…16 Microfluidic methods use an outer continuous phase to focus the flow of an inner prepolymer phase from which the spheres are fabricated. 17 Once droplets are produced they can be solidified using appropriate reactions. UV photopolymerisation is an effective and convenient way to crosslink droplets produced by microfluidic methods using functionalised polymers, [18][19][20] however, limited studies have been performed with hydrogels 21 and fewer again with synthetic or biosynthetic hydrogels.…”

Single-cell technologies in environmental omics

“…Bacterial, yeast, plant, insect, and mammalian cells can be analyzed using microdroplet technology on agarose (Zhang et al, 2012a) or other microgels (Rossow et al, 2012). Even multicellular organisms can be encapsulated and grown in a droplet (Clausell-Tormos et al, 2008).…”