We describe a perfusion-based microfluidic device for three-dimensional (3D) dynamic primary human hepatocyte cell culture. The microfluidic device was used to promote and maintain 3D tissue-like cellular morphology and cell-specific functionality of primary human hepatocytes by restoring membrane polarity and hepatocyte transport function in vitro without the addition of biological or synthetic matrices or coagulants. A unique feature of our dynamic cell culture device is the creation of a microenvironment, without the addition of biological or synthetic matrices or coagulants, that promotes the 3D organization of hepatocytes into cord-like structures that exhibit functional membrane polarity as evidenced by the expression of gap junctions and the formation of an extended, functionally active, bile canalicular network.

We present a poly(dimethylsiloxane) (PDMS)-based hot embossing process for low-cost rapid prototyping of plastic microfluidic devices. Unlike conventional hot embossing process, the process presented here uses a 2 mm thick PDMS mold fabricated via soft lithography, two ¾" wide binder clips, two standard 1 mm thick 1" × 3" microscope glass slides and an oven. Micro-scale features were successfully replicated in 1.5 mm thick polystyrene slides from various PDMS molds. Also, PDMS molds can be reused for many replications without any damages.

Many biological materials and cell substrates are very soft (Young's modulus < 500 Pa) and it is difficult to characterize their mechanical properties. Here we report local elasticity of the surface layers of Matrigel™ films used for cell culture. We used a new measurement technology, Mechanical Imaging Interferometry, to obtain point mechanical measurements over micron-sized areas. The median values of 650 Pa +/-400 Pa (# measurements, n=50), determined by the Hertz contact model, agree well with bulk measurements, however on the micro-scale the films were heterogeneous and contained regions distinctly stiffer than average (1-2 kPa). The first measurement of yield strengths of 170 Pa +/-100 Pa (n=43) indicate that Matrigel™ films deform plastically at stress levels of similar scale to cell tractional forces.