RESUMO
Perfusion of porous scaffolds transports cells to the surface to yield cellular constructs for 3D models of disease and for tissue engineering applications. While ceramic scaffolds mimic the structure and composition of trabecular bone, their opacity and tortuous pores limit the penetration of light into the interior. Scaffolds that are both perfusable and amenable to fluorescence microscopy are therefore needed to visualize the spatiotemporal dynamics of cells in the bone microenvironment. In this study, a hybrid injection molding approach was designed to enable rapid prototyping of collector arrays with variable configurations that are amenable to longitudinal imaging of attached human mesenchymal stem cells (hMSCs) using fluorescence microscopy. Cylindrical collectors were arranged in an array that is permeable to perfusion in the xy-plane and to light in the z-direction for imaging from below. The effects of the collector radius, number, and spacing on the collection efficiency of perfused hMSCs was simulated using computational fluid dynamics (CFD) and measured experimentally using fluorescence microscopy. The effect of collector diameter on simulated and experimental cell collection efficiencies followed a trend similar to that predicted by interception theory corrected for intermolecular and hydrodynamic forces for the arrays with constant collector spacing. In contrast, arrays designed with constant collector number yielded collection efficiencies that poorly fit the trend with collector radius predicted by interception theory. CFD simulations of collection efficiency agreed with experimental measurements within a factor of two. These findings highlight the utility of CFD simulations and hybrid injection molding for rapid prototyping of collector arrays to optimize the longitudinal imaging of cells without the need for expensive and time-consuming tooling.
RESUMO
Fabrication of microfluidic devices by photolithography generally requires specialized training and access to a cleanroom. As an alternative, 3D printing enables cost-effective fabrication of microdevices with complex features that would be suitable for many biomedical applications. However, commonly used resins are cytotoxic and unsuitable for devices involving cells. Furthermore, 3D prints are generally refractory to elastomer polymerization such that they cannot be used as master molds for fabricating devices from polymers (e.g. polydimethylsiloxane, or PDMS). Different post-print treatment strategies, such as heat curing, ultraviolet light exposure, and coating with silanes, have been explored to overcome these obstacles, but none have proven universally effective. Here, we show that deposition of a thin layer of parylene, a polymer commonly used for medical device applications, renders 3D prints biocompatible and allows them to be used as master molds for elastomeric device fabrication. When placed in culture dishes containing human neurons, regardless of resin type, uncoated 3D prints leached toxic material to yield complete cell death within 48 hours, whereas cells exhibited uniform viability and healthy morphology out to 21 days if the prints were coated with parylene. Diverse PDMS devices of different shapes and sizes were easily cast from parylene-coated 3D printed molds without any visible defects. As a proof-of-concept, we rapid prototyped and tested different types of PDMS devices, including triple chamber perfusion chips, droplet generators, and microwells. Overall, we suggest that the simplicity and reproducibility of this technique will make it attractive for fabricating traditional microdevices and rapid prototyping new designs. In particular, by minimizing user intervention on the fabrication and post-print treatment steps, our strategy could help make microfluidics more accessible to the biomedical research community.