Surface functionalized iron oxide nanoparticles for applications in biomedical sciences
Abstract
This thesis focuses on the functionalization of iron oxide nanoparticles (Fe3O4) and their
applications in biomedical sciences. Each chapter represents an independent research project that
has been conducted within a different collaboration. For each project, magnetic Fe3O4 iron oxide
nanoparticles have been functionalized or modified to suit its requirements. This thesis aims to
show how versatile and most promising Fe3O4 iron oxide nanoparticles are, and how their unique
properties in size, shape, and magnetism can be utilized for a broad range of applications in
biomedicines.
Chapter 2 focuses on magnetic resonance (MR), computed tomography (CT), and intravascular
ultrasound (IVUS) as essential diagnostic imaging techniques and how iron oxide nanoparticles
can potentially be used as contrast agents across all three imaging modalities. Contrast agents are
commonly used to enhance the imaging quality and thus provide more detail for assessment.
However, previous studies using nanoparticles for MR and CT were prepared with surface coating
stabilizers, which in turn can compromise the use in clinical studies. In this chapter, gold-iron
oxide nanoparticles (Au*MNP) are presented as a multi-modal contrast agent. Using a chemically
grafting method without stabilizers, presenting nanoparticles with pristine surfaces that allow for
further functionalization in molecularly targeted theragnostic applications.
In Chapter 3, the response of HepaRG liver cells to nanoparticles is examined in two methods, 2D
and 3D cell culturing. By analyzing the cell response in 2D and 3D cultures an accurate estimation
of the toxicity of nanoparticles can be made. The toxicity of iron oxide was assessed using
commercially available cell assays (CellTiter-Glo and PrestoBlue), however, the experiments
suggested some restrictions that could alter the data and therefore resemble inaccurate results. For
this purpose, non-invasive imaging techniques based on impedance (xCELLigence system) and
Coherent Anti-Stokes Raman Scattering (CARS) were used to analyze the cell toxicity. The results
showed that those methods provided a much deeper insight into the cell viability and proliferation
of HepaRG cells. Furthermore, valuable data on the immediate effects in real-time and long-term
exposures can be captured of the same culture.
Chapter 4 presents the cell internalization process of iron oxide nanoparticles, captured using the
unique holographic cell imaging technique of a HoloMonitor M4 microscope. In most cases where
the cell internalization process is monitored, only one or two cells can be visualized and tracked at
the same time. This is not the case with this technique, where hundreds of cells can be
simultaneously visualized, analyzed, and monitored over time. Unlike single-cell observation, the
system takes pictures of the cell culture at a high capture rate, allowing to observe and interpret
the cell dynamics, cell morphologies, and cell reactions to nanoparticles (e.g., toxicity and
apoptosis). Measuring the kurtosis and skewness of MCF-7 cancer cells for 72 hours after
nanoparticle exposure, showed that cell splitting and proliferation took place, and no extraordinary damages or cell death was caused by the internalization of the particles. Furthermore, every step
of the internalization process was monitored and captured in visible data for the first time.
Chapter 5 demonstrates magnetic molecularly imprinted polymer networks and spheres (MMIPs)
for the selective binding of antibiotics. MIPs are polymers that can be synthesized with highly
selective and reusable binding sites. Their combination with magnetic iron oxide can be used as a
useful tool to monitor and remove antibiotic pollutants from freshwater sources and food products.
MMIPs are prepared using a microemulsion technique containing vinyl-functionalized iron oxide
to selectively bind the model antibiotics erythromycin (ERY) and ciprofloxacin (CPX). The results
show that MMIPs prepared using this technique are highly selective towards their respective
template molecule in methanol/water and milk matrix, are recyclable, and most importantly open
to modification.
In Chapter 6, zebrafish larvae are exposed to polyethylene glycol coated iron oxide supported gold
nanoparticles for further toxicity assessment. In general, toxicological data is gathered in vivo,
however, the translational values from in vitro to in vivo are sometimes questionable due to the
complexity of the organism. Zebrafish larvae are used as an intermediate method for in vivo
experiments, as they can be used 96 hours after hatching, unlike larger animals such as mice, rats,
or rabbits which take a much longer time to reach adulthood, sometimes up to 3 months. Also, a
single female zebrafish can spawn about 200-300 eggs per week, which can generate an extensive
data set from a small-scale experimental setup. The results presented in this chapter showed a
100% survival rate of all exposed zebrafish larvae between a range of concentrations from 0 - 2mM
(0 - 463.1 mg/L).
Chapter 7 summarizes the key findings and developments presented in this thesis, suggestions for
future work within each research project, and proposes future applications.
Chapter 8 includes a list of journal publications produced from this thesis.