Recently, there has been considerable interest in the use of microfluidics to miniaturize assays and control cellular microenvironments in cell-based studies. We fabricated a microfluidic system in which cells attach only at a central region along the microchannel, using which we assessed cellular dynamic responses to a concentration gradient of biological factors (Fig. 1). In conventional methods, cells randomly attach to the entire inner surface of the microchannel, making it difficult to precisely characterize cellular responses and migration activities. Our system, however, consists of three parts: a branched channel, a main channel for the cell culture, and two side channels flowing into the main channel. The branched channel is designed to generate a stable concentration gradient by mixing and dividing two external inputs into six discrete streams. The main channel is coated with a cell-repellent cross-linked albumin. A laminar flow of polyethyleneimine (PEI) generated with the main and two side channels is used to change specific regions in the main channel from being cell-repellent to cell-adhesive. In this scheme, fibroblasts or hepatocytes are attached to the central region along the main channel. Then the remaining surface is changed from being cell-repellent to cell-adhesive with PEI, thereby facilitating cell migration from the first site in response to a concentration gradient. With this system, we have conducted cytotoxic assays with anticancer agents and surfactants, as well as assays of migration across a concentration gradient of a biological factor. Thus, this microfluidic system, combined with our cell micropatterning technology, may be a useful tool in fabricating controlled cell microenvironments for fundamental biological studies and tissue engineering applications.

Fig. 1 Microfluidic cell culture device with concentration gradient generator.