Search

We report an in situ method for three-dimensionally resolved temperature measurement in microsystems. The temperature of the surrounding fluid is correlated from Brownian diffusion of suspended nanoparticles. We use video-microscopy in combination with image analysis software to selectively track nanoparticles in the focal plane. This method is superior with regards to reproducibility and reduced systematic errors since measuring Brownian diffusivity does not rely on fluorescence intensity or lifetime of fluorophores. The efficacy of the method is demonstrated by measuring spatial temperature profiles in various microfluidic devices that generate temperature gradients and by comparing these results with numerical simulations. We show that the method is accurate and can be used to extract spatial temperature variations in three dimensions. Compared to conventional methods that require expensive multiphoton optical sectioning setups, this technique is simple and inexpensive. In addition, we demonstrate the capability of this method as an in situ tool for simultaneously observing live cells under the microscope and monitoring the local temperature of the cell medium without biochemical interference, which is crucial for quantitative studies of cells in microfluidic devices.

We report a microfabricated field flow fractionation device for continuous separation of subcellular organelles by isoelectric focusing. The microdevice provides fast separation in very small samples while avoiding large voltages and heating effects typically associated with conventional electrophoresis-based devices. The basis of the separation is the presence of membrane proteins that give rise to the effective isoelectric points of the organelles. Simulations of isoelectric focusing of mitochondria in microchannels are used to assess design parameters, such as dimensions and time scales. In addition, a model of Joule heating effects in the microdevice during operation indicates that there is no significant heating, even without active cooling. The device is fabricated using a combination of photolithography, thin-film metal deposition/patterning, and electroplating techniques. We demonstrate that in the microfluidic devices, mitochondria from cultured cells migrate under the influence of an electric field into a focused band in less than 6 min, consistent with model predictions. We also illustrate separation of mitochondria from whole cells and nuclei as well as the separation of two mitochondrial subpopulations. When automated and operated in parallel, these microdevices should facilitate high-throughput analysis in studies requiring separation of organelles.

We describe the design, construction, and characterization of microfluidic devices for studying cell adhesion and cell mechanics. The method offers multiple advantages over previous approaches, including a wide range of distractive forces, high-throughput performance, simplicity in experimental setup and control, and potential for integration with other microanalytic modules. By manipulating the geometry and surface chemistry of the microdevices, we are able to vary the shear force and the biochemistry during an experiment. The dynamics of cell detachment under different conditions can be captured simultaneously using time-lapse videomicroscopy. We demonstrate assessment of cell adhesion to fibronectin-coated substrates as a function of the shear stress or fibronectin concentration in microchannels. Furthermore, a combined perfusion-shear device is designed to maintain cell viability for long-term culture as well as to introduce exogenous reagents for biochemical studies of cell adhesion regulation. In agreement with established literature, we show that fibroblasts cultured in the combined device reduced their adhesion strength to the substrate in response to epidermal growth factor stimulation.

We report fabrication, characterization, and use of microfluidic analysis devices containing surface-immobilized cell-capturing molecules. Amino-terminated biotin ligands are immobilized onto the luminal surface of a microdevice and effectively support serf-assembly of proteins, antibodies, and mammalian cells. For this purpose, chemical vapor deposition (CVD) polymerization is used to functionalize PDMS-made microfluidic devices with poly-[para-xylylene carboxylic acid pentafluorophenolester-copara-xylylene]. The resulting reactive coating shows excellent adhesion when deposited in thin films (~100 nm, and the distribution of the pentafluorophenol ester groups is reasonably uniform within the microchannel inner surface, as examined by fluorescence microscopy. The utility of these devices for cell-based bioassays is demonstrated by monitoring the concentration-dependent effect of the disintegrin echistatin on cell adhesion. The described assay format could be relevant to clinical research in various fields, including angiogenesis research.