A microperfused incubator for tissue mimetic 3D cultures |
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Authors: | Jelena Vukasinovic D Kacy Cullen Michelle C LaPlaca Ari Glezer |
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Institution: | (1) Woodruff School of Mechanical Engineering, Georgia Institute of Technology, 771 Ferst Drive, Atlanta, GA 30332-0405, USA;(2) Coulter Department of Biomedical Engineering, Georgia Institute of Technology, 313 Ferst Drive, Atlanta, GA 30332-0535, USA |
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Abstract: | High density, three-dimensional (3D) cultures present physical similarities to in vivo tissue and are invaluable tools for pre-clinical therapeutic discoveries and development of tissue engineered constructs.
Unfortunately, the use of dense cultures is hindered by intra-culture transport limits allowing just a few layer thick cultures
for reproducible studies. In order to overcome diffusion limits in intra-culture nutrient and gas availability, a simple scalable
microfluidic perfusion platform was developed and validated. A novel perfusion approach maintained laminar flow of nutrients
through the culture to meet metabolic need, while removing depleted medium and catabolites. Velocity distributions and 3D
flow patterns were measured using microscopic particle image velocimetry. The effectiveness of forced convection laminar perfusion
was confirmed by culturing 700 μm thick neural-astrocytic (1:1) constructs at cell density approaching that of the brain (50,000
cells/mm3). At the optimized flow rate of the nutrient medium, the culture viability reached 90% through the full construct thickness
at 2 days of perfusion while unperfused controls exhibited widespread cell death. The membrane aerated perfusion platform
was integrated within a miniature, imaging accessible enclosure enabling temperature and gas control of the culture environment.
Temperature measurements demonstrated fast feedback response to environmental changes resulting in the maintenance of the
physiological temperature within 37 ± 0.2°C. Reproducible culturing of tissue equivalents within dynamically controlled environments
will provide higher fidelity to in vivo function in an in vitro accessible format for cell-based assays and regenerative medicine. |
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