Microfluidic devices for automated cell synchronization and analysis

A detailed understanding of the mechanisms by which bacteria adhere to surfaces is essential to elucidate how biofilms form and bacteria infect. The ability to track these adhesion events at the single cell level with fluorescence microscopy provides insight into the adhesion process and heterogeneity of the process within a given cell population. To study bacterial adhesion with improved temporal precision, we developed microfluidic devices for the synchronization and analysis of individual bacteria. With microfluidic devices, we are able to precisely control the local environment in which the cells reside and to monitor the behavior of individual bacteria.;In our initial work, we used microfluidic devices with single channels to study permanent adhesion events of the bacterium Caulobacter crescentus. From video data that tracked movement and adhesion of individual cells to the microchannel surface, we determined the timescale for holdfast production (∼20 s) in wild-type cells and compared these adhesion events to bacteria without pili. To expand our measurement capabilities, we moved the cell synchronization step on chip and developed a microfluidic "baby machine." The microfluidic devices have integrated pumps and valves to control the movement of cells and media. Synchronized populations are collected from the device at intervals as short as 10 min and at any time over four days. We also monitored the length of the stalk organelle in response to changes in the phosphate concentration of the environment. Our on-chip synchronization method overcomes limitations with conventional physical cell separation methods that consume large volumes of media, require manual manipulations, have lengthy incubation times, are limited to one collection, and lack precise temporal control of collection times. We now have a closed, automated system that streamlines the steps of cell seeding, culture, synchronization, and analysis.