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One major goal of biology is to provide a quantitative description of cellular physiology. This task is complicated by population effects, which perturb culture conditions and mask the behavior of the individual cell. To overcome these limitations, the construction and operation of a microfluidic bioreactor is presented. The new reactor concept guarantees constant environmental conditions and single cell resolution, thus it was named Envirostat (environment, constant). In the Envirostat, cells are contactless trapped by negative dielectrophoresis (nDEP) and cultivated in a constant medium flow. To control chip temperature, a Peltier device was constructed. Joule heating by nDEP was quantified with Rhodamine B in dependence of applied voltage, field mode, medium conductivity, and flow velocity. The integration of the Joule heating effect in the temperature control allowed setting and maintaining the cultivation temperature. For single cell cultivation of Saccharomyces cerevisiae, medium composition changes below 0.001% were estimated by computational fluid dynamic simulation. These changes were considered not to influence cell physiology. Finally, single S. cerevisiae cells were cultivated for more than four generations in the Envirostat, thus showing the applicability of the new reactor concept. The Envirostat facilitates single cell research and might simplify the investigation of hitherto difficult to access biological phenomena such as the true regulatory and physiological response to genetic and environmental perturbations.

Single cell analysis is mainly limited to single time-point measurements, without the possibility to track behavioral changes of a single cell or its descendents. Here, the integration of a spatiotemporal single cell lab-on-a-chip system with an automated cultivation device allows single cell analysis under defined growth conditions and, especially, semiautomated cell retrieval and growth kinetic analysis of the single cell descendants. Performance of the new platform was evaluated using the yeast Saccharomyces cerevisiae. The yeast was singularized in the lab-on-a-chip (Envirostat), which combines the possibility of cell cultivation with cell analysis. Singularized cells were collected in a microtiter plate and cultivated in a semiautomated cultivation device (Bioscreen C). S. cerevisiae showed highly reproducible and glucose concentration independent growth kinetics in population experiments. Yet, growth kinetics in cultures inoculated with only one or few cells exhibited strong variations, because of an unexpected growth phenotype: colony formation in submerse cultures. Interestingly, the colony-like structures grew for more then 60 h and were stable for at least 82 h, despite rigorous shaking. Cell agglomeration due to pseudohyphal growth could be excluded, suggesting changes in cell wall properties of yeast populations starting from a single yeast. Single cell analysis still exhibits unexpected obstacles. Nevertheless, this new single cell analysis platform can be used for studying cellular dynamics of single cells and expanded cell populations thereof. '