Metal ion sensors using Tunable Resistive Pulse Sensing

There is a drive to develop rapid, portable and simple methods for detecting heavy metal ions. Due to their toxic nature, heavy metal ions are monitored in aqueous solutions such as drinking water. Standard methods for metal detection rely on instrumentation such as atomic absorption/emission and mass spectrometry. These are often costly and do not allow for rapid on-site or real-time measurements. The aim of this PhD was to develop and optimise tunable resistive pulse sensing (TRPS) for sensing metal ions. This combines nanomaterials, dual molecular recognition with an emerging nanopore technology. TRPS is a label-free portable sensor that allows characterisation of particles based on their size, concentration and charge. Monitoring changes upon the particle surface via changes to the particle charge could be a powerful analytical tool for studying metal ion binding and new sensors. Tuning functional groups on the nanoparticle surface will allow for an array of metal ions to be detected. Nanoparticles will be modified with functional groups that bind to metal ions in solution, in turn this will change the charge on the nanoparticle which will be studied using TRPS. Particle velocity through the pore is dependent on particle charge so changes on the nanoparticle surface can be monitored. The literature review in Chapter 1 focuses on the use of different ligands for the detection of metals focusing on aptamers and modified nanoparticles. The application of the theory of resistive pulse sensors (RPS), which is the main sensing platform within the thesis is covered in detail however these sensors to date have little use in metal ion detection. The theory behind RPS follows the literature review. This covers the theory of transport through a conical nanopore, a brief introduction to zeta potential and particle surface charge and ion current rectification. Before developing a metal ion sensor, the translocation of a particle through the pore, focusing on its relative velocity needed to be understood. Chapter 3 demonstrates how changes in the double layer can affect the measured particle velocity. Understanding how the double layer changes with ionic strength and pH is essential in designing a metal ion sensor where the velocity of the particle through the pore is being measured. The work presented in Chapter 3 gave confidence that TRPS could be used to monitor metal ion binding to the surface of nanoparticles. The nanoparticles were modified with a ligand (APTES) and DNA. The subsequent particle velocities differ to those of the unmodified particles, making TRPS a suitable platform for monitoring changes upon a nanoparticle surface. Building on the knowledge gained from Chapter 3, particle translocation velocities were used for the detection of copper (II) on the surface of modified nanoparticles, Chapter 4. Changes in particle velocity through the nanopore allows for detection of copper (II) as low as 1 ppm and at 10 ppm with competing metal ions present. Chapter 4 also presents the first use of studying pulse waveshape for the detection of an analyte. At low ionic strengths, particles passing through the conical pore generated a biphasic pulse containing a conductive pulse and resistive pulse. The biphasic pulse behaviour was used to monitor changes on the nanoparticle surface, and infer the presence of ions within the particles double layer. The method can be easily adapted to different analytes by altering the ligand used. As an alternative to a particle-based assay, a pore-based assay was developed which exploited the current rectification properties of the conical pores used in TRPS. Chapter 5 presents the use of Layer-by-Layer (LbL) assembly of polyelectrolytes onto the surface of the polyurethane pore for the modification of the pore wall, a DNA aptamer was then easily immobilized onto the pore wall. Vascular Endothelial Growth Factor (VEGF) was chosen as the analyte prior to developing a metal ion assay as it was a system studied in more detail in the literature and within the group. An advantage of TRPS is the particle-by-particle analysis. This allows for simple multiplex detection by using particles of two different sizes to detect two different analytes. In Chapter 6 the methodology and techniques from Chapter 4 is applied to the multiplexed detection of lead (II) and mercury (II) using particle translocation velocities to detect the metal ion binding to DNA aptamers. The method is applicable over a large range of ionic strengths with little interference from a high salt content. Finally, to advance the multiplexed concept, the two independent aptamer sequences used in Chapter 6 are merged together. While both aptamer halves retain their initial functionality and bind to the respective metals, the location of the binding and change in DNA structure with respect to the particles surface is the dominating factor in determining the sensitivity of the RPS technology