Fine temporal control of the medium gas content and acidity and on-chip generation of series of oxygen concentrations for cell cultures.
We describe the design, operation, and applications of two microfluidic devices that generate series of concentrations of oxygen, [O(2)], by on-chip gas mixing. Both devices are made of polydimethylsiloxane (PDMS) and have two layers of channels, the flow layer and the gas layer. By using in-situ measurements of [O(2)] with an oxygen-sensitive fluorescent dye, we show that gas diffusion through PDMS leads to equilibration of [O(2)] in an aqueous solution in the flow layer with [O(2)] in a gas injected into the gas layer on a time scale of approximately 1 sec. Injection of carbon dioxide into the gas layer causes the pH in the flow layer to drop within approximately 0.5 sec. Gas-mixing channel networks of both devices generate series of 9 gas mixtures with different [O(2)] from two gases fed to the inlets, thus creating regions with 9 different [O(2)] in the flow layer. The first device generates nitrogen-oxygen mixtures with [O(2)] varying linearly between 0 and 100%. The second device generates nitrogen-air mixtures with [O(2)] varying exponentially between 0 and 20.9%. The flow layers of the devices are designed for culturing bacteria in semi-permeable microchambers, and the second device is used to measure growth curves of E. coli colonies at 9 different [O(2)] in a single experiment. The cell division rates at [O(2)] of 0, 0.2, and 0.5% are found to be significantly different, further validating the capacity of the device to set [O(2)] in the flow layer with high precision and resolution. The degree of control of [O(2)] achieved in the devices and the robustness with respect to oxygen consumption due to respiration would be difficult to match in a traditional large-scale culture. The proposed devices and technology can be used in research on bacteria and yeast under microaerobic conditions and on mammalian cells under hypoxia.
Polinkovsky
Mark
M
Department of Physics, University of California, San Diego, , 9500 Gilman Drive, MC 0374, La Jolla, CA 92093, USA.
Gutierrez
Edgar
E
Levchenko
Andre
A
Groisman
Alex
A
D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods; D003470:Culture Media; D004129:Dimethylpolysiloxanes / Q000737:chemistry; D004867:Equipment Design; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development; D006863:Hydrogen-Ion Concentration; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods; D009584:Nitrogen / Q000032:analysis; D010100:Oxygen / Q000032:analysis*; D012996:Solutions
D003470:Culture Media; D004129:Dimethylpolysiloxanes; D012996:Solutions; C013830:baysilon; D009584:Nitrogen; D010100:Oxygen