Building integrated cell-based microsystems : fabrication methodologies, metrology tools and impact on cellular physiology

Abstract

This thesis presents the development of new tools for the realization and characterization of complex, integrated biological microsystems. These tools involve - (1) the implementation and characterization of a new material system for biological microsystems, and its application to the design of a single-bioparticle trap, (2) the creation of a lipid vesicle-based metrology tool for the characterization of dielectrophoresis-based microsystems, and (3) the construction of live-cell stress reporters for the characterization of electric-field effects on cell physiology in microsystems. Materials for biological microsystems have been dominated by the elastomer polydimethylsiloxane (PDMS) and the photopolymer SU-8 which form the core of soft lithography based fabrication. The design of highly integrated microsystems in which microfluidic conduits are coupled with electrical manipulation techniques, however, is difficult to realize with soft lithography. This thesis presents a new fabrication technique based on photopatternable silicones. I show that these photopatterned silicones are able to generate free-standing structures, exhibit low autofluorescence, enable alignment with pre-patterned substrates, and are biocompatible. These unique properties enable the generation of a new type of single-particle trap, which would be challenging to realize using traditional techniques. This new material system is now well poised to enable the design and construction of new biological microsystems that could not be previously realized. In addition to new material systems and fabrication technology, designing highly integrated cell-based biological microsystems requires the use of synthetic, cell-like particles.

(cont.) To date, polystyrene microspheres have served as surrogates for cells in characterizing these systems, despite the fact that they serve as poor models of cells. This thesis presents a new metrology tool for the characterization of microsystems, based on phopsphohlipid vesicles. I show the ability to modulate the electrical properties of such vesicles and generate electrically addressable vesicles and for use in the characterization of dielectrophoretic-based microsystems. Finally, as biological microsystems gain in complexity, the need to characterize their impact on living cells is of paramount importance. This thesis presents the construction of stress reporting cell lines that form the core of live-cell metrology tool for assaying physiological impact in Microsystems that utilize electric fields to manipulate cells. Specifically, the response of these stress reporter cells to conditions typically experienced in a dielectrophoretic trap are explored over a wide range of voltages, frequencies and durations. The results obtained point to the role of multiple stressing agents and provide new insight in to stress initiation across frequency. The use of such sensors is now well poised to study the physiological impact of microsystems across a wide range of conditions. Together, the tools presented in this thesis promise to enable the development of systems with unprecedented flexibility of design as well as functionality.