| The patch-clamp technique is a powerful tool for the investigation of transmembrane ionic currents and the gating nature of the membrane proteins that control them. The patch-clamp technique has been used extensively to study many electrophysiological cell properties, such as the characterization of the kinetics and steady-state effects of toxins, chemical agonists, antagonists, and drugs in ion channels, transporters, and pumps. Although the traditional methods used to perform the patch-clamp technique have been very successful, conventional pipettes do not satisfy the needs of all applications for which the patch-clamp technique could be used. Examples of electrophysiological applications that would benefit from uniquely tailored patch-clamp systems include measuring dose-response curves for single cells, high-throughput screening, applications needing optical access to the cell (e.g., confocal and fluorescence microscopy), and applications needing rapid intracellular and extracellular perfusion.;This dissertation describes the design, fabrication, and testing of a planar patch-clamp system that addresses some of the limitations present in traditional patch-clamp systems. In the presented system, a planar patch-clamp substrate is integrated with microfluidic components that replace the patch pipette and preparation bath in a traditional patch-clamp system. A four-channel software-controlled pressure-control system was developed to control the flow of solutions in the microfluidic components.;The planar patch-clamp substrates have been tested and are shown to form high-resistance seals in excess of 1 GO using Chinese hamster ovary (CHO-K1) and mouse erythroleukemia (MEL) cell lines. Testing shows that the microfluidic components and pressure control system are able to drive MEL cells to patch apertures with a flow velocity of 100 mum/s, trap cells from the suspension onto patch apertures, and exchange the extracellular fluid environment. |
