Nonequilibrium nanoparticle dynamics for the development of Magnetic Particle Imaging

Thesis (Ph.D.)--University of Washington, 2019Magnetic Particle Imaging (MPI) is a promising new medical imaging platform currently in the preclinical stage. MPI uses iron oxide nanoparticles as tracers, and exploits their nonlinear magnetization response to external oscillating magnetic fields. The nanoparticle response is directly measured to create an image in MPI, and so optimizing nanoparticle properties as well as external magnetic field conditions in order to obtain improved image signal is crucial to the development of MPI as a clinical platform. A deep understanding of the physics of nanoparticle relaxation is fundamental to optimization and further development of MPI. This work focuses on modeling the nonequilibrium dynamics of magnetic nanoparticles in order to optimize conditions for MPI and develop new therapeutic and diagnostic functionalities that can be integrated with MPI. A summary of theoretical models of nanoparticle dynamics is presented, and computational nonequilibrium models are outlined, which currently represent the most sophisticated methods for modeling nanoparticle dynamics. These models are verified and supported through experiment. Using these models, nanoparticle relaxation is explored in depth; the effect of applied field amplitude and frequency, as well as nanoparticle size, on the resulting relaxation mechanism and timescale is investigated in detail. These insights are then applied to the optimization of drive field conditions and nanoparticle size for MPI image resolution and sensitivity. A core size of 28 nm is found to be optimal, with additional tuning required according to specific field conditions used. Finally, a procedure for multicolor MPI, in which the signal from nanoparticles of different types or in different environments is separated, is developed. A multi-channel image reconstruction approach is outlined, and discrimination based on nanoparticle core size is demonstrated, resulting in successful generation of a multicolor MPI image of a 2D phantom. A procedure for quantitative temperature estimation with MPI is also proposed and verified experimentally. Overall, this work provides a theoretical foundation for nanoparticle relaxation physics in MPI, enabling further development towards the clinical stage

Comparative Modeling of Frequency Mixing Measurements of Magnetic Nanoparticles Using Micromagnetic Simulations and Langevin Theory

Dual frequency magnetic excitation of magnetic nanoparticles (MNP) enables enhanced biosensing applications. This was studied from an experimental and theoretical perspective: nonlinear sum-frequency components of MNP exposed to dual-frequency magnetic excitation were measured as a function of static magnetic offset field. The Langevin model in thermodynamic equilibrium was fitted to the experimental data to derive parameters of the lognormal core size distribution. These parameters were subsequently used as inputs for micromagnetic Monte-Carlo (MC)-simulations. From the hysteresis loops obtained from MC-simulations, sum-frequency components were numerically demodulated and compared with both experiment and Langevin model predictions. From the latter, we derived that approximately 90% of the frequency mixing magnetic response signal is generated by the largest 10% of MNP. We therefore suggest that small particles do not contribute to the frequency mixing signal, which is supported by MC-simulation results. Both theoretical approaches describe the experimental signal shapes well, but with notable differences between experiment and micromagnetic simulations. These deviations could result from Brownian relaxations which are, albeit experimentally inhibited, included in MC-simulation, or (yet unconsidered) cluster-effects of MNP, or inaccurately derived input for MC-simulations, because the largest particles dominate the experimental signal but concurrently do not fulfill the precondition of thermodynamic equilibrium required by Langevin theory

Nanopore-Based Conformational Analysis of a Viral RNA Drug Target

Nanopores are single-molecule sensors that show exceptional promise as a biomolecular analysis tool by enabling label-free detection of small amounts of sample. In this paper, we demonstrate that nanopores are capable of detecting the conformation of an antiviral RNA drug target. The hepatitis C virus uses an internal ribosome entry site (IRES) motif in order to initiate translation by docking to ribosomes in its host cell. The IRES is therefore a viable and important drug target. Drug-induced changes to the conformation of the HCV IRES motif, from a bent to a straight conformation, have been shown to inhibit HCV replication. However, there is presently no straightforward method to analyze the effect of candidate small-molecule drugs on the RNA conformation. In this paper, we show that RNA translocation dynamics through a 3 nm diameter nanopore is conformation-sensitive by demonstrating a difference in transport times between bent and straight conformations of a short viral RNA motif. Detection is possible because bent RNA is stalled in the 3 nm pore, resulting in longer molecular dwell times than straight RNA. Control experiments show that binding of a weaker drug does not produce a conformational change, as consistent with independent fluorescence measurements. Nanopore measurements of RNA conformation can thus be useful for probing the structure of various RNA motifs, as well as structural changes to the RNA upon small-molecule binding

A structural model for microtubule minus-end recognition and protection by CAMSAP proteins.

CAMSAP and Patronin family members regulate microtubule minus-end stability and localization and thus organize noncentrosomal microtubule networks, which are essential for cell division, polarization and differentiation. Here, we found that the CAMSAP C-terminal CKK domain is widely present among eukaryotes and autonomously recognizes microtubule minus ends. Through a combination of structural approaches, we uncovered how mammalian CKK binds between two tubulin dimers at the interprotofilament interface on the outer microtubule surface. In vitro reconstitution assays combined with high-resolution fluorescence microscopy and cryo-electron tomography suggested that CKK preferentially associates with the transition zone between curved protofilaments and the regular microtubule lattice. We propose that minus-end-specific features of the interprotofilament interface at this site serve as the basis for CKK's minus-end preference. The steric clash between microtubule-bound CKK and kinesin motors explains how CKK protects microtubule minus ends against kinesin-13-induced depolymerization and thus controls the stability of free microtubule minus ends