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Evaporation is a critical problem when handling submicroliter volumes of fluids. This paper characterizes this problem as it applies to microfluidic cell culture in poly(dimethylsiloxane) (PDMS) devices and provides a practical solution. Evaporation-mediated osmolality shifts through PDMS membranes with varying thicknesses (10, 1, 0.2, or 0.1 mm) were measured over 96 h. Even in humidified cell culture incubators, evaporation through PDMS and associated shifts in the osmolality of culture media was significant and prevented mouse embryo and human endothelial cell growth and development. A simple diffusion model, where the measured diffusion coefficient for PDMS matches reported values of ~[10.sup.-9] [m.sup.2]/s, accounts for these evaporation and osmolality shifts. To overcome this problem, a PDMS-parylene-PDMS hybrid membrane was developed that greatly suppresses evaporation and osmolality shifts, yet possesses thinness and the flexibility necessary to interface with deformation-based microfluidic actuation systems, maintains the clarity for optical microscopy, and enables the successful development of single-cell mouse embryos into blastocysts under static conditions and culture of human endothelial cells under dynamic recirculation of submicroliter volumes of media. These insights and methods demonstrated specifically for embryo and endothelial cell studies will be generally useful for understanding and overcoming evaporation-associated effects in microfluidic cell cultures.

Endothelial cells (ECs) lining the inner lumen of blood vessels are continuously subjected to hemodynamic shear stress, which is known to modify EC morphology and biological activity. This paper describes a self-contained microcirculatory EC culture system that efficiently studies such effects of shear stress on EC alignment and elongation in vitro. The culture system is composed of elastomeric microfluidic cell sheafing chambers interfaced with computer-controlled movement of piezoelectric pins on a refreshable Braille display. The flow rate is varied by design of channels that allow for movement of different volumes of fluid per variable-speed pump stroke. The integrated microfluidic valving and pumping system allowed primary EC seeding and differential sheafing in multiple compartments to be performed on a single chip. The microfluidic flows caused ECs to align and elongate significantly in the direction of flow according to their exposed levels of shear stress. This microfluidic system overcomes the small flow rates and the inefficiencies of previously described microfluidic and macroscopic systems respectively to conveniently perform parallel studies of EC response to shear stress.

Evaporation is a critical problem when handling submicroliter volumes of fluids. This paper characterizes this problem as it applies to microfluidic cell culture in poly- (dimethylsioxane) (PDMS) devices and provides a practical solution. Evaporation-mediated osmolality shifts through PDMS membranes with varying thicknesses (10, 1, 0.2, or 0.1 mm) were measured over 96 h. Even in humidified cell culture incubators, evaporation through PDMS and associated shifts in the osmolality of culture media was significant and prevented mouse embryo and human endothelial cell growth and development. A simple diffusion model, where the measured diffusion coefficient for PDMS matches reported values of ~10-9 m²/s, accounts for these evaporation and osmolality shifts. To overcome this problem, a PDMS-parylene-PDMS hybrid membrane was developed that greatly suppresses evaporation and osmolality shifts, yet possesses thinness and the flexibility necessary to interface with deformation-based microfluidic actuation systems, maintains the clarity for optical microscopy, and enables the successful development of single-cell mouse embryos into blastocysts under static conditions and culture of human endothelial cells under dynamic recirculation of submicroliter volumes of media. These insights and methods demonstrated specifically for embryo and endothelial cell studies will be generally useful for understanding and overcoming evaporation-associated effects in microfluidic cell cultures. [ABSTRACT FROM AUTHOR]