A Microfluidic Device for Physical Extraction of Intracellular Proteins from Bacterial Cells

The analysis of intracellular proteins from bacterial cells typically requires a cell population of ~1000 or more due to the small cell size. Effective combination of concentration and lysis steps is desired for such analysis to be carried out on a microfluidic platform. In here, we report a simple microfluidic device that integrates the capture of the bacterial cells using a microscale bead array and the rapid electrical lysis for the release of intracellular proteins. We study the retention of Escherichia coli cells with different concentrations in this type of bead array and the optimal electrical parameters for the electroporative release of intracellular proteins. The bead array can be replaced after each run and this allows the device to be used for multiple times. Our design provides a simple solution to the extraction of intracellular proteins from a bacterial cell population based entirely on physical methods without applying chemical or biological reagents. Our device forms a critical basis for bacterial proteomic studies based on microfluidics. Introduction There are numerous assays applied in biological research that are essentially based on analysis of intracellular materials from cells. Conventional assays based on nucleic acids or proteins from cells require at least hundreds to thousands of cells as the starting material. Recently new tools such as capillary electrophoresis and microfluidics have allowed researchers to analyze intracellular contents from a low number of cells . Due to powerful amplification techniques such as polymerase chain reaction and detection techniques such as laser-induced fluorescence, the analysis of intracellular nucleic acids and proteins at the single cell level has become routine for mammalian cells . However, the analysis of intracellular materials from bacterial cells, especially proteins, still has to be conducted based on a fairly large number of cells. Bacteria are important model organisms to study regulatory networks and protein functions because they have relatively small genomes and low number of protein components compared to eukaryotic cells . Furthermore, there have been established methods for genetic manipulation of some bacteria which make the studies of protein functions attainable. Finally, bacterial proteomic studies provide crucial information on the adaptation networks that are important for bacterial survival and such knowledge is critical for the discovery of antibacterial drugs 15, . For example, a single Escherichia coli cell has a length of 2 μm and a diameter of 0.8 μm which yield a volume of ~1 fl . This volume is about three orders of magnitude smaller than that of a typical eukaryotic somatic cell. Analysis of intracellular proteins from bacterial cells typically requires