Microfluidics for cell and system biology

Microfluidics is an active and quickly developing area of applied science and engineering. It can be defined as construction and testing of devices and arrangements, where liquids flow through systems of microscopic channels, usually 1-100 microns thick and 10-1000 microns broad. It has numerous current and even more potential applications, both industrial and scientific, in fine chemistry, biochemistry, biology and medicine. Among the advantages and applications of microfluidics are the following.
" The microfluidic chemical reactors are of very small size, which leads to dramatic reduction in required amounts of expensive biochemical reagents. The system can be closed and compartmentalized, or opened and mixed, either actively or diffusively.
" Individual microscopic devices can be integrated in complex and composite arrangements, so that potentially a whole biochemical laboratory may be constructed on a single microchannel chip.
" The microchannels are easily optically accessible. Chemical and biological processes, single cells and macromolecules can be observed under a high magnification microscope in real time.
" Flow in the microchannels is always laminar and can be arranged quite precisely. Therefore, chemical composition of the medium and flow conditions in the microchannels can be well controlled with high resolution in space and time.
We are building microfluidic devices out of PDMS, which is rubber-like transparent silicon elastomer, using soft lithography. We have got a small facility in the lab (expected to be completely operational by November 2002), where we do the whole process of microfabrication. Elasticity of the silicon rubber and its good adhesion to glass allows for a fast and inexpensive fabrication process. It also allows construction of multilayer devices with flexible membranes between the layers. The membranes can bent and locally block flow in a channel in one layer, when excessive pressure is applied to a crossing channel in another layer. So that a membrane can act as a microscopic valve (typically 100x100 microns in size), which can be opened or closed, and allow or stop the flow within a few milliseconds. Precise tuning of pressures driving the flows (typically with a resolution of 1 Pa) gives us a tool for an accurate control of flow conditions.
We have developed microfluidic devices for different projects in collaboration with a few groups specializing in cell and system biology. The first microfluidic device creates two narrow (about 3 microns wide) submerged parallel streams of chemicals, which can be switched on and off independently with about 10 ms resolution in time. The distance between the steams is controlled with a micron resolution, and can be varied from virtually zero to a few hundreds microns. So, two sides of a single eukaryotic cell (10-20 microns in diameter) attached to a substrate can be independently and sharply exposed to different (or the same) chemicals. This way reaction of the cell to signals coming from different sides can be studied. It is relevant to the issue of chemotaxis of amoebas and neutrophils in particular, and to the problem of intracellular resolution of signaling networks, pathways and events in general.
Another project is on chemotropism of yeasts. The haploid yeast cells have an ability to mate under some conditions. The individual cells are known to find their mating partners by gradient of a pheromone, the mating -factor, and get extended in the direction of the partner. We immobilize the freely floating yeast cells in a flow and expose them to a well defined gradient of the pheromone. Varying parameters of the gradient field in space and time, we are trying to obtain quantitative data on the chemotropic reactions of the yeasts.
Still another project is building a microfluidic biostat for bacteria. It is a device, where (highly mobile) bacterial cells are trapped in a chamber, which is impenetrable for them, but well penetrable for chemicals. By permanently exchanging the medium using either molecular diffusion or active flow, we can keep the chemical contents of the chamber constant. This way we hope to keep the bacteria in an exponential growth phase up to very high cell densities, which may be hard to reach otherwise. This high density combined with the exponential growth may well correspond to physiological conditions for some bacteria. On the other hand, bacteria in a flask slow down their division rate when they reach sufficiently high density, because of decreasing concentration of nutrients and increasing concentration of impeding metabolites. Observing the bacterial cultures in the micro-chamber under a high magnification microscope should give new information about their behavior and mutual influence at high densities.

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