High Fidelity Photopatterning and Characterization of Hydrogels for Three-Dimensional Cell Co-Culture

Abstract

Hydrogels provide a unique environment for the in-vitro study of cell behavior in 3D, and the technique of photolithography allows for controlled cell patterning by enabling the creation of well-defined hydrogel structures. However, current photolithographic techniques are limited in their ability to produce hydrogels on the micro scale with thicknesses sufficient for use in a true 3D cell culture system. We demonstrate methods for patterning hydrogels derived from both biological and synthetic polymers with enhanced fidelity and thickness. Oligo(poly(ethylene glycol) fumarate) (OPF) and chondroitin sulfate methacrylate (CSMA) polymers were synthesized using previously described methods1,2. OPF and CSMA hydrogels 550 - 3000 μm in size were photocrosslinked in microfluidic devices purged with nitrogen gas under 365 nm UV light for 12 minutes at an intensity of 10.5 mW/cm2. Hydrogels photopatterned under nitrogen were found to have significantly higher thickness than those patterned in the presence of room air, exceeding 1 mm for gels with widths greater than 600 - 800 μm. Hydrogel size correlated with the size of the photomask, enabling facile system calibration. The photopatterning methods were also employed to laminate two differentially swollen hydrogels to form a single construct with a well-defined and stable interface visualized using fluorescence microscopy. These results demonstrate that we are capable of producing hydrogels with improved spatial resolution at the μm to mm scale that may be laminated to form well defined patterns of cells in a 3D co-culture system. References: 1Jo S et al. Macromolecules. 2001;34:2839-44. 2Bryant S et al. Macromolecules. 2004; 37:6726-6733.