Cellular metabolomics can provide critical information regarding the effects of diseased states. Traditional cell culture and experimentation is done in dishes or flasks that are in static environments, but this limits the biological comparison since flow affects cell morphology and function. Microfluidics provides a biomimetic solution with small culture channels that enable flow and have dimensions on the same order of magnitude as blood vessels. Microfluidics also provides a solution to the issues with conventional cell lysing such as experimenter variation and the involved scraping and centrifugation process. Our system uses a combination of lysing solvent, electroporation, and a Peltier cooler with lysing time under 5 seconds and a temperature of -5°C that provides optimal enzymatic quenching.
Figure 1: Cell Culture, Lysis, and Extraction Setup. Cells are cultured in chips for ~48hrs before lysing. The flow selector actuates to rinse cells with water to remove salt before the lysing buffer is applied. Lysates are collected and stored at -80°C.
Figure 2: Incorporation of Peltier Cooler. Set-up identical to Figure 1, but upon actuation of the lysing solvent the chip is placed on a chilled Peltier cooler that brings the average temperature of the chip down to -5 °C to reduce enzymatic activity.
Figure 3: Capture of Mammalian cells on a microfluidic platform, using photopolymerized monolith. critical to understanding cell-cell interactions in diabetic complications is the separation and metabolic analysis of each cell type after the interaction. seen above are mammalian cells stained with calcein AM which have been captured on-chip using a photopolymerized monolith.
Figure 4. platform for macrophage-EC co-culture experiment. A. Configuration for charging sample loop with macrophage cell suspension. Note a second microfluidic device for rapid and automated trapping and lysis of macrophages. B. Macrophage suspension introduced to media loop and cycled via reciprocating pump. C. peristaltic pump is turned off. Water is flushed through the system, rinsing the EC and pushing the macrophages onto a photopolymerized frit in the channel of the second device. D. Using two syringes, lysing solvent is simultaneously introduced to the microfluidic devices, and high voltage is applied to the EC culturing device. The second 6-port valve directs the lysate to collect. E. A high voltage relay switch directs the high voltage to the second device, and the macrophage lysate is collected.