Rapid Prototyping of Multilayer Microphysiological Systems

Abstract

Here we report benchtop fabrication of multilayer thermoplastic organs-on-chips via laser cut and assembly of double sided adhesives. Biocompatibility was evaluated with Caco-2 cells and primary human intestinal organoids. Chips with Luer fluidic interfaces were economical ($2 per chip) and were fabricated in just hours without use of specialized bonding techniques. Compared with control static Transwell™ cultures, Caco-2 and organoids cultured on chips formed confluent monolayers expressing tight junctions with low permeability. Caco-2 cells on chip differentiated ~4 times faster compared to controls and produced mucus. To demonstrate the robustness of laser cut and assembly, we fabricated a dual membrane, tri-layer gut chip integrating 2D monolayers, 3D cell culture, and a basal flow chamber. As proof of concept, we co-cultured a human, differentiated monolayer and intact organoids in a chip with multi-layered contacting compartments. The epithelium exhibited 3D tissue structure and organoids formed in close proximity to the adjacent monolayer. The favorable features of thermoplastics, such as low gas and water vapor permeability, in addition to rapid, facile, and economical fabrication of multilayered devices, make laser cut and assembly an ideal fabrication technique for developing organs-on-chips and studying multicellular tissues.

Reconfigurable Microphysiological Systems for Modeling Innervation and Multitissue Interactions

“…GelPins are thin layers of material within our custom microfluidic devices that facilitate the formation of discrete, yet contiguous structures in either the x-y and/or z planes ( Figure 1A). This manufacturing technique, in combination with photocrosslinkable hydrogels, enables rapid (hours) and inexpensive (<$2 in materials per MPS) prototyping of custom, [5] multilayer, 3D MPS with nearly limitless geometric configuration possibilities ( Figure 1B-D). As a proof of concept, GelPins were leveraged to establish the first biomimetic in vitro model of the cardiac sympathetic nervous system (SNS) for investigating the effects of sympathetic activity and increase beat rate on cardiomyocytes (CMs) and innervation, respectively.…”

“…Device Fabrication: Organ-chip devices were fabricated using a modified approach to previously described methodology. [5] A laser engraver system (Epilog Zing 16, Epilog Laser) cut and shaped flow paths in double-sided adhesive tape (966 adhesive, 3 m), polymethyl methacrylate (PMMA, McMaster-Carr) and polyethylene terephthalate (PET, McMaster-Carr) sheets, and a polycarbonate (PC) track etched membrane (GE Healthcare) using material specific speed and power settings ( Figure 1B). Each individual layer was then assembled layer-bylayer into a complete device with the aid of precut alignment holes and a custom alignment jig.…”

“…[4,8] Toward overcoming these limitations, we have previously reported on the development of a novel "cut and assemble" manufacturing technique as an alternative to traditionally used, soft-lithography fabrication approaches ( Figure 1B). [5] While continuing to improve in resolution, laser engravers are not able to fabricate freestanding features such as microtunnels and/or micropost to enable robust compartmentalization as their silicon counterparts. Therefore, we developed and integrated micrometer-scale GelPins within our devices to enable establishment of temporary liquid-air interfaces, that once polymerized, formed discrete hydrogel regions ( Figure 1B,C).…”

“…Based on each MPS design's unique geometry and flow rate parameters, using this simulation, fluidic shear stresses may be calculated and adjusted to promote endothelial monolayer formation. [5] However, while membranes seeded with endothelial cells may be used to help mimic vasculature-like diffusion, membranes fail to fully recapitulate the tissue-vessel iterations and act as additional diffusion barriers.…”

“…[4] Toward overcoming these limitations, our previously reported "cut and assemble" manufacturing technique was expanded to exploit the meniscus pinning effect via GelPins to compartmentalize 3D cell-laden materials within a tailorable MPS. [5]…”