We describe the design, operation, and applications of two microfluidic devices that generate series of concentrations of oxygen, [O2], 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 [O2] with an oxygen-sensitive fluorescent dye, we show that gas diffusion through PDMS leads to equilibration of [O2] in an aqueous solution in the flow layer with [O2] in a gas injected into the gas layer on a time scale of ∼1 sec. Injection of carbon dioxide into the gas layer causes the pH in the flow layer to drop within ∼0.5 sec. Gas-mixing channel networks of both devices generate series of 9 gas mixtures with different [O2] from two gases fed to the inlets, thus creating regions with 9 different [O2] in the flow layer. The first device generates nitrogen-oxygen mixtures with [O2] varying linearly between 0 and 100%. The second device generates nitrogen-air mixtures with [O2] 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 [O2] in a single experiment. The cell division rates at [O2] of 0, 0.2, and 0.5% are found to be significantly different, further validating the capacity of the device to set [O2] in the flow layer with high precision and resolution. The degree of control of [O2] 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.
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