A Lab-on-PCB biomedical diagnostic platform for on-chip protein preconcentration.

: (Alternative Format Thesis)

Abstract

Non-communicable diseases account for just under three quarters of deaths worldwide, with low- and middle-income countries accounting for over three quarters of these. This percentage, and number, is continuing to rise year after year, and with it the importance of monitoring these conditions. The ability to diagnose, monitor and react to changes is vital in the fight against these diseases. However, due to lack of resources, it is not always possible to bring the patients in need to a dedicated facility. Point-of-Care testing is one way of ensuring that a greater proportion of people in need are monitored, but issues with accessibility and financing remain a critical hurdle. Lab-on-a-Chip technology takes laboratory scale processes and shrinks them into portable devices on the scale of a credit card which require little to no specialist training. A sub-class of this technology that uses polymer based printed circuit boards, PCBs, is Lab-on-PCB which allows for mass manufacturing and the associated reduction in costs.

Whilst these miniaturised devices solve many problems and allow for testing to be taken to, and potentially operated by, the patient, the reduced sample volumes needed to run the assays bring their own problems. This project aimed to work to address this problem. By increasing the concentration of a charged target analyte, here proteins were the focus, strain is lessened on the development of the assay and specifically on the limit of detection for the device. The device developed here utilised printed circuit boards, to create a module to preconcentrate proteins that would be integrable with other Lab-on-a-Chip sensors and assays. This module incorporated the electrochemical control of pH with optical methods to develop a device capable of performing microfluidic isoelectric focusing on an individually addressed PCB array.

The first experiments were focused on the use of thermally evaporated gold electrodes on glass slides. Optical methods were established, and the characterisation and optimisation of a fluorescent dye was carried out before the development of the electrochemical system. Self-assembled monolayers were formed on the gold electrodes, utilising the passive formation of well-ordered surface functionalisation using thiol terminated molecules. Electrochemical polymerisation was then introduced to the surface chemistry to improve the electrochemical reversibility of the oxidation and reduction reactions of the monolayer. Studies into the behaviour of the functionalised electrodes were then performed to identify and test the control over the acidification of the environment local to the electrodes. These experiments were run in tandem with simulations in COMSOL Multiphysics, however, due to financial restrictions, the results from these analyses were not furthered.

Transference of the methods developed in the first experimental chapter on glass to a PCB platform were then carried out. Cleaning and functionalisation of gold electrodes on the PCB was explored and custom PCB arrays were designed, fit for purpose. An assessment of the new platform using the same fluorescent dye was completed and the optimal potential to electrochemically change the pH over time was investigated. Further characterisation was carried out and the spatial resolution of the device was identified which guided the final design of the PCB array for the purpose of conducting experiments to preconcentrate proteins.

The final experimental chapter details the development of the open-air device, consisting of the custom PCB array and holder, to control multiple electrodes simultaneously and trial preconcentration of bovine haemoglobin, a protein with properties compatible with the restraints of the dye used. These experiments showcased the promise of the device, and the incorporation of microfluidic channels was successfully realised after initial problems with dye concentration and optical path length through the channels were resolved. Preconcentration experiments were detailed using two proteins and two methods of assessing the concentration of these target molecules were devised.

In short, this thesis summarises the development of a Lab-on-PCB module designed to preconcentrate proteins using the electrochemical generation of acid through the application of a potential to a polymerised self-assembled monolayer. The functionality of the device was assessed optically using a fluorescent dye and the progression from glass to PCB to microfluidics is detailed alongside the cleaning and optimisation processes. This is the first report of protein preconcentration on a lab-on-a-PCB with protein preconcentration achieved in two different formats; showing a quantitative increase in concentration of more than 3-fold in one configuration and qualitative results for its microfluidic form. Ultimately the device is shown to have promise, demonstrating the clear preconcentration of at least one protein in solution and the proposed future work is outlined.

Date of Award	24 Apr 2024
Original language	English
Awarding Institution	University of Bath
Supervisor	Despina Moschou (Supervisor) & Pedro Estrela (Supervisor)