High-throughput microfluidic platforms applied to immunoassay-based diagnostics and environmental monitoring

The research presented in this thesis is motivated by the aim to employ microfluidic devices for high-throughput analysis. Two applications, in particular, have been explored: diagnostics and environmental sensing. Miniaturization and multiplexing of laboratory procedures leads to a decrease of the cost and response time of the test. Moreover miniaturization allows the system to become portable, enabling use in the field without bulky external equipment. This need is paving the way for innovative approaches and microfluidics can be a suitable substitute for standard bench top techniques. In order to be appealing, these new technologies should have the same or even better performances over the standard techniques, not only in terms of time and cost, but also quality of results. First, we developed a multilayer microfluidic platform for high-throughput immunoassay analysis. The device is composed of 384 units, each of them containing 4 mechanically induced trapping of molecular interactions (MITOMI) buttons, where the interaction of antibodies and antigens occurs. Therefore, the platform allows us to perform 1'536 tests, and we show that it can be useful for large scale screening in order to identify functional antibody pairs. However this system has two limitations: the experimental time (~10 hours) and the sensitivity (pM). In the second part, we presented another device modified in order to overcome the aforementioned drawbacks and to become more suitable for point of care applications. This new version of the device has 16 independent units, and the MITOMI buttons are patterned with femtoliter wells for digital analysis. We show how digital ELISA improves the sensitivity and allows us to detect down to fM concentrations in human serum. We also show how the combination of both digital and analog detection can broaden the dynamic range. A portable automatic controller has also been built to control the valves and the flow throughout the chip and a USB microscope has been exploited in order to measure the fluorescent signal. In the last part we present a different application for high-throughput microfluidic-based devices. A microfluidic device composed of 769 units has been developed for bacterial cell culture and environmental monitoring. Each unit can be considered as a pixel and the entire device as a biodisplay. Each unit contains a specific bacterial strain, which is deposited using microspotting techniques. We show how different bacterial strains can be cultured over time without contamination between pixels and how they can be monitored after induction. Cells can be spotted according to a specific pattern on the chip and express fluorescent protein reporters in the presence of specific molecules allowing an easy to interpret result of the test. For example, the image of a “skull and crossbones” appears if the analyzed sample is contaminated with arsenic. Overall, the presented work provides several microfluidic platforms for high-throughput analysis of clinical or environmental samples; these devices are useful tools for the study of a large number of proteins and molecular diagnostic applications and to characterize bacterial strains and environmental monitoring.