Development of a Separation-Based Sensor using Microdialysis Coupled to Microchip Electrophoresis with Electrochemical Detection for Monitoring Catecholamines

Abstract

Microdialysis is a powerful separation technique capable of simultaneously monitoring multiple analytes in the extracellular fluid of the brain. This technique generates small sample volumes in a continuous flow stream. Traditional methods used for sample analysis forfeit temporal information regarding dynamic neurochemical processes due to the larger volumes necessary for analysis. Additionally, sample acquisition methods traditionally involve some form of tethering or anesthetizing the animal under study, greatly reducing the available behavioral information. In order to preserve both temporal resolution and behavioral information, the ideal analysis system is one that can be employed on-line, has fast analysis times of small sample volumes, and can be placed on a freely-roaming animal. Microdialysis sampling coupled on-line to microchip electrophoresis with electrochemical detection creates a separation-based sensor that fulfills these constraints. The ability to place the device directly on-animal, without tethering, allows for the neurochemical information to be correlated with the animal’s behavior, allowing for further understanding of the neurochemical basis behind each behavior. Additionally, neuroactive drug metabolism can be monitored alongside behavior when employing an on-animal separation-based sensor, potentially aiding in drug development. The goal of this thesis is therefore to develop a separation-based sensor that is capable of monitoring neurochemicals in vivo. Towards this aim, the separation and detection of analytes in the dopamine metabolic pathway was accomplished using microchip electrophoresis with electrochemical detection at a carbon electrode. The substrate material in this separation was also optimized. In order to integrate this separation and detection with microdialysis sampling, a novel fabrication procedure was developed. This procedure creates a PDMS/glass hybrid device capable of integrating hydrodynamic microdialysis flow with electrophoretic flow and detection at a carbon electrode using a flow-gated interface. Lastly, the developed method was used to monitor the dopamine metabolic pathway in vivo in rat after the administration of L-DOPA. In the future, the complete device and associated instrumentation can be used remotely and on-animal, for near-real time in vivo monitoring.