Design And Validation Of A Microfluidic Chip For 3D Cell Culture

Microfluidic chips offer a number of advantages,such as low consumption of reagents,ease of precise control of the fluid flow,and ease of mimicking the cellular microenvironment in vitro,and they have been widely used in the field of cell analysis.Cell culture is the foundation of various of cell analyses.In this paper,A microfluidic chip with a micropillar array chamber for three-dimensional(3D)cell culture was designed and validated.The chip consisted of a polydimethylsiloxane(PDMS)channel plate and a glass cover plate.One cell culture chamber composed of two rows of micropillar arrays and two lateral channels for transporting medium were integrated on the PDMS channel plate.The spacing between micropillars directly affects the performance of the chip,which is critical for the design of the chip.It needs to not only ensure that the mixture of cells and extracellular matrix mimics can be steadily injected in the chamber,but also meet that nutrients in the medium and the cell metabolites can quickly diffuse.First of all,based on computational fluid dynamics,the injection process of the mixture of cells and collagen and the diffusion process of nutrients in the medium were numerically simulated separately in this paper.A 2D geometric model used for simulating the injection process of the mixture was constructed by using the two-phase level set mode of COSMOL Multiphysics software.The injection process of the collagen solution was influenced by the injection pressure,the capillary force and the surface tension.In the chamber,the pressure and the capillary force played major roles and the collagen solution flowed rapidly.However,the collagen solution flowed much more slowly between micropillars because it was hindered by the viscous force and the surface tension.As the spacing increased,the flow hindrance decreased gradually,and the collagen solution had a tendency to break through the micropillars and enter the lateral channel.When the spacing increased to 100 ?m,the collagen solution entered the lateral channel after 200 ms.Similarly,A 3D geometric model used for simulating the diffusion process of nutrients in the medium was constructed by using the species transport mode of ANSYS Fluent software.The simulation results indicated the nutrients flowed quickly in the lateral channel,and meanwhile it diffused into the chamber through micropillar arrays.The spacing has an effect on the diffusion rate of nutrients in the medium.The larger the spacing,the faster the diffusion rate.The diffusion equilibrium was reached after 220 s when the spacing was 50 ?m.Based on the above simulation results,and taking into account the fabrication difficulties and the performances of the chip,50 ?m was chosen as the optimal spacing.Then,the optimized microfluidic chip was fabricated based on the simulation results,and the injection process of the mixture and the diffusion process of nutrients were validated by experiments.The experimental results indicate that the optimized spacing by numerical simulation meets the requirements of the microfluidic chip.On the one hand,the mixture of cells and extracellular matrix mimics can be steadily injected in the chamber,and the leakage of the mixture into the lateral channel can be effectively blocked.On the other hand,nutrients in the medium can quickly diffuse into the chamber.Finally,neural stem cells were 3D cultured in the optimized chip,and the concentrations of glucose and lactic acid were continually measured for a long time.The experimental results demonstrate that the microenvironment constructed in the chip has an excellent stability and is capable of long-term 3D cell culture,and has a great potential for a variety of cell analyses.