D. Yang, A. Jennings, J. Männik
University of Tennessee,
Keywords: self-organizing process, microfluidics, pressure actuated microvalve, nanofabrication
Summary:Cells maintain their structure and reproduce through numerous self-organizing processes. The common approach to understand how cellular organization is maintained is to perturb this organization and then observe a response from the cell. Essentially all approaches used by cell biologists in these studies enable the assessment of only ensemble-averaged responses, overlooking significant cell-to-cell variations that exist in a population. The current methods are also not suitable to study processes in cells with fast response times. Here, we describe the development of two microfluidic platforms that allow mechanical and chemical perturbation of cellular organization while following the cellular response at single cell level in real time using a high resolution fluorescence microscope. The platform designed for studies involving chemical perturbation consists of PDMS channels with lateral dimensions comparable to the diameter of a bacterial cell. Each channel connects to a larger flow channel allowing for controlled and rapid introduction of different chemical agents to the growth medium. While the fluidic portion of the platform functions as expected, our studies show that cells have growth limitations in the channels. We will discuss the origin of this limitation. The platform designed for studies involving mechanical perturbation consists of pressure actuated microvalves. Bacteria are placed under a valve that when closed leads to bacterial deformation and shape changes. As we have shown before a long term deformation of bacterial cell leads to drastic changes in its morphology [2, 3]. The platform enables to systematically study the effect of cell shape on cellular function. The two platforms present promising complementary approaches to probe, in a quantitative and systematic way, processes that maintain cellular functions. In addition to bacterial cells, these two platforms can also be useful in studies of yeast and other single-celled organisms. References 1. Characterization of small microfluidic valves for studies of mechanical properties of bacteria, D. Yang, C. M. Greer, B. P. Jones, A. D. Jennings, S. T. Retterer, and J. Männik, J. Vac. Sci. Technol. B 33, (2015) 06F202. 2. Robustness and accuracy of cell division in Escherichia coli in diverse cell shapes, J. Männik, F. Wu, F. J. H. Hol, P. Bisicchia, D. Sherratt, J. E. Keymer and C. Dekker, Proc. Natl. Acad. Sci. U. S. A. 109 (2012) 6957. 3. Bacterial growth and motility in sub-micron constrictions, J. Männik, R. Driessen, J. E. Keymer and C. Dekker, Proc. Natl. Acad. Sci. U. S. A. 106 (2009) 14861.