Flow analysis on microcasting with degassed polydimethylsiloxane micro-channels for cell patterning with cross-linked albumin.

Yigang Shen,Takayuki Kawai,Nobuyuki Tanaka,Hidenori Nagai,Yo Tanaka,Hironori Yamazoe,Shunsuke Furutani,Talib Al-Ameri

doi:10.1371/journal.pone.0232518

Abstract

Patterned cell culturing is one of the most useful techniques for understanding the interaction between geometric conditions surrounding cells and their behaviors. The authors previously proposed a simple method for cell patterning with an agarose gel microstructure fabricated by microcasting with a degassed polydimethylsiloxane (PDMS) mold. Although the vacuum pressure produced from the degassed PDMS can drive a highly viscous agarose solution, the influence of solution viscosity on the casting process is unknown. This study investigated the influences of micro-channel dimensions or solution viscosity on the flow of the solution in a micro-channel of a PDMS mold by both experiments and numerical simulation. It was found experimentally that the degassed PDMS mold was able to drive a solution with a viscosity under 575 mPa·s. A simulation model was developed which can well estimate the flow rate in various dimensions of micro-channels. Cross-linked albumin has low viscosity (1 mPa·s) in aqueous solution and can undergo a one-way dehydration process from solution to solid that produces cellular repellency after dehydration. A microstructure of cross-linked albumin was fabricated on a cell culture dish by the microcasting method. After cells were seeded and cultivated on the cell culture dish with the microstructure for 7 days, the cellular pattern of mouse skeletal myoblast cell line C2C12 was observed. The microcasting with cross-linked albumin solution enables preparation of patterned cell culture systems more quickly in comparison with the previous agarose gel casting, which requires a gelation process before the dehydration process.

Highlights

In the field of cell biology, because the spatial arrangement of cells is an important factor in morphogenesis and tissue functions [1,2], artificially generated spatial patterns of cells have been often used
A PDMS mold with the designed structure was fabricated by the photolithography method (Fig 2A) and cross-linked albumin supplied at the open ends of the micro-channels was draw into the micro-channels
This was assumed to be derived from the meniscus between the side wall of the PDMS mold and bottom surface, Microcasting flow analysis for cell patterning by cross-linked albumin which was formed during the dehydration process

Summary

Introduction

In the field of cell biology, because the spatial arrangement of cells is an important factor in morphogenesis and tissue functions [1,2], artificially generated spatial patterns of cells have been often used. The authors have succeeded in patterning cells using a microstructure cast of agarose, one of the non-cell-adhesive substances [14,15] In these two studies, a casting solution was driven into a degassed polydimethylsiloxane (PDMS) mold by the accumulated vacuum pressure in the PDMS. It is known that when a high viscosity solution flows in a narrow channel, it generates a large pressure loss, reducing the flowability This might cause the introduction of solution to take longer than expected from vacuum production time constant, resulting in insufficient casting of the microstructure. To clarify the durability of crosslinked albumin under cell culture conditions, this study compared the cell patterns of mouse skeletal myoblasts C2C12 with microstructures made of native and cross-linked albumin. This study compared the cell patterns of mouse skeletal myoblasts C2C12 with microstructures made of native and cross-linked albumin

Materials and methods

Results and discussion

Conclusion