A High-Throughput Method for In Vitro Generation and Studies of OxygenMicrogradients.

Oxygen;Micro Gradient;Microfluidics;Metabolic;Mass Transfer;MEMS;Electrical Engineering;Engineering;Electrical Engineering

Pinelis, MikhailNajafi, Khalil ;

University of Michigan

关键词: Oxygen;    Micro Gradient;    Microfluidics;    Metabolic;    Mass Transfer;    MEMS;    Electrical Engineering;    Engineering;    Electrical Engineering;   

Others  :  https://deepblue.lib.umich.edu/bitstream/handle/2027.42/75849/pinelis_1.pdf?sequence=1&isAllowed=y

瑞士|英语

来源: The Illinois Digital Environment for Access to Learning and Scholarship

【 摘 要 】

A high-throughput method for in vitro generation of oxygen and mass transfer microgradients was developed. This method not only overcomes many limitations of the previous approaches, but also provides a comprehensive experimental and modeling framework for future studies.To demonstrate this method, microfluidic devices and associated fabrication processes were developed.The microfluidic devices consisted of capillary channels with sizes ranging from 30-300 μm in height to 0.3-3 mm in width, loading reservoirs and microfluidic interconnects.The overall device size was 38×18×1.3 mm3.The fabrication process was based on SU-8 photolithography and glass to SU-8 bonding. Because of the sizes of the capillaries, diffusion, and not convection, dominated mass transfer and oxygen microgradients were generated, by diffusion limitations and cellular respiration, within 0.5–3 mm of the capillary edges.The method is based on diffusion and, therefore, more closely mimics the in vivo microenvironments within multicellular tissues.Measurements of cell viability, pH, differentiation and oxygen were performed for C2C12 and HeLa cells cultured in the mass transfer gradients.Oxygen was measured using the fluorescence lifetime imaging method (FLIM) with spatial resolution of 1 mm and measurement resolution of 0.1 parts per million of dissolved oxygen.This method lends itself to high-throughput experimentation; as many as 30 capillary experiments were run in a 24-hour period including cell loading, gradient formation and imaging.Observable differences in cell morphology became apparent 12-24 hours after seeding.The developed finite element model (FEM) accommodates a wide range of device geometries and metabolic parameters and couples cellular metabolism with diffusion effects.The model predictions for dissolved oxygen levels and live cell densities were within 5-15% of measured data.