MULTIPLEXED SINGLE CELL ANALYSIS OF ENZYME ACTIVITY AND GENE EXPRESSION FOR CAPILLARY ELECTROPHORESIS

Chemical cytometry is a valuable single cell platform that has been used to study a wide variety of biochemicals inside a cell. However, chemical cytometry assays can be limited in scope often relying on probes or analyzing chemically similar molecules. Single cell sequencing is a relatively new platform that has a broad scope capable of measuring thousands of genes but is generally limited to studying nucleic acids. A combined platform that combines the assay flexibility from chemical cytometry with the broad scope of single cell sequencing would be a valuable tool for single-cell analysis. This dissertation discusses the development of generalized capillary electrophoresis (CE) based chemical cytometry method and instrumentation that incorporates single-cell nucleic acid collections during the chemical cytometry assay. A modular and adaptable automated CE instrument and software was developed in chapter 2. Additionally, the performance of an open source low-cost fluorescence detector for CE is given in chapter 3. Finally, a general method for nucleic acids is presented in chapter 4.Doctor of Philosoph

Control strategy for a flexible analytical chemistry robotics system

This thesis is the result of work carried out during more than two years on a Teaching Company Scheme. Liaison took place between Rhone-Poulenc Agriculture Limited (the industrial partner), hereafter referred to as RPAL or the company, and Middlesex University (the academic partner). The aim of the Scheme was to realise the design, development, commissioning, testing and validation of an intelligent robotic system for sample analysis of trace pesticides and metabolites in order to enable quicker product development. Due to the complexity of the project and the range of technical expertise and skills needed for its implementation, three associates participated in the Programme. I joined as the second associate. With my degree in Industrial Engineering, I have been in overall charge of developing the computational aspects of the system, from control overview to implementation and validation. Two distinct types of studies will be carried out with the robot based system: • Routine extraction of pesticide from soil or plant material, which is compound as well as analyst dependant. • Method development studies, to optimise those routine extraction processes. Traditional strategies of control were not applicable for such system because we were dealing with the automation of a non repetitive process involving non-deterministic operations (evaporation, filtration, etc.). The resulting control system should provide a high degree of flexibility to allow workcell reconfiguration without involving any reprogramming. Modularity is also a must if expansion and upgrading to new technologies and equipment is to involve relatively little cost and effort. In addition, all generated data has to be stored and reported following Good Laboratory Practice (GLP) standards. As the system is both large and flexible in operation, it has proven a real challenge to develop. Software had to be written that can - among its many tasks - allow unrestricted analyst choice, optimise system performance, detect, prioritise and act upon error signals, dynamically schedule robot and instrument operation in real time, trace samples as they pass through the system and generate results as reports stored in databases. The system is now virtually complete, and is presently undergoing the last stages of the validation. Due to the success of this scheme, further cooperative ventures are being planned between Rhone-Poulenc and Middlesex University in both the UK and France